Near infrared microbial elimination laser systems (nimels)

ABSTRACT

Methods, systems, and apparatus for Near Infrared Microbial Elimination Laser Systems (NIMELS) including use with medical devices are disclosed. The medical devices can be situated in vivo. Suitable medical devices include catheters, stents, artificial joints, and the like. NIMELS methods, systems, and apparatus can apply near infrared radiant energy of certain wavelengths and dosimetries capable of impairing biological contaminants without intolerable risks and/or adverse effects to biological moieties other than a targeted biological contaminant associated with traditional approaches described in the art (e.g., loss of viability, or thermolysis). Lasers including diode lasers may be used for one or more light sources. A delivery assembly can be used to deliver the optical radiation produced by the source(s) produced to an application region that can include patient tissue. Exemplary embodiments utilize light in a range of 850 nm-900 nm and/or 905 nm-945 nm at suitable NIMELS dosimetries.

RELATED APPLICATIONS

This application is a continuation of International Application No.PCT/US2006/030434 filed 3 Aug. 2006, which claimed the benefit of U.S.Provisional Application Ser. No. 60/705,630, filed 3 Aug. 2005; and acontinuation-in-part of International Application No. PCT/US2006/028616filed 21 Jul. 2006, which claimed priority to U.S. Provisional PatentApplication Ser. No. 60/701,896, filed Jul. 21, 2005; U.S. ProvisionalPatent Application Ser. No. 60/711,091, filed Aug. 23, 2005; U.S.Provisional Patent Application Ser. No. 60/780,998, filed Mar. 9, 2006;and U.S. Provisional Patent Application Ser. No. 60/789,090, filed Apr.4, 2006; this application is also a continuation-in-part of U.S.application Ser. No. 10/776,106 filed 11 Feb. 2004, which is acontinuation-in-part of U.S. application Ser. No. 10/406,493 filed 28Aug. 2006, which claimed priority to U.S. Provisional Patent ApplicationNo. 60/406,493 filed 28 Aug. 2002; the contents of all of whichapplications are incorporated herein by reference in their entireties.

This application is also related to U.S. Provisional Application Ser.No. 60/705,630, filed 3 Aug. 2005, entitled “Near Infrared MicrobialElimination Laser (NIMEL) System and Devices Based Thereon,” thecontents of which are incorporated herein in their entirety byreference, and which is assigned to the assignee of the presentapplication. This application is also related to the followingapplications, of common assignee as the present application: NearInfrared Microbial Elimination Laser (NIMEL) System,” U.S. ProvisionalPatent Application Ser. No. 60/701,896, filed 21 Jul. 2005; “NearInfrared Microbial Elimination Laser (NIMEL) System,” U.S. ProvisionalPatent Infrared Microbial Elimination Laser (NIMEL) System,” U.S.Provisional Patent Application Ser. No. 60/711,091, filed 23 Aug. 2005;“Method and Apparatus for the Treatment of, and Prevention of Recurrenceof Finger and Toenail Infections,” U.S. Provisional Patent ApplicationSer. No. 60/780,998, filed 9 Mar. 2006; and “Method and Device for theUniform Illumination of NIMELS Optical Energy and Dosimetry to aBiological Containment in a Biological Moiety,” U.S. Provisional PatentApplication Ser. No. 60/789,090, filed 4 Apr. 2006; all of whichapplications are incorporated herein in their entirety by reference.

BACKGROUND

1. Field of the Disclosure

The present disclosure relates to methods, systems, and apparatus forselectively reducing the level of a biological contaminant in a targetsite, including target sites encompassing or partially including one ormore medical devices. The present disclosure also encompassestherapeutic modalities, and more particularly, relates to methods,devices, and systems using optical radiation.

2. Background of the Disclosure

Several E. coli species and other enterococci are known to haveintrinsic and acquired resistance to most antibiotics making themsignificant nosocomial pathogens in human and animal disease. Boyce, etal., J. Clin. Microbiol. 32(5):1148-53 (1994); Donskey, et al., N. Engl.J. Med. 343(26):1925-32 (2000); Landman, et al., J. Antimicrob.Chemother. 40(2):161-70 (1997). Human infections that are caused byenterococci can include endocarditis, bacteremia, urinary tractinfection, wound infection, and intra-abdominal and pelvic infections.

For a great number of these infections, the organisms originate from thepatient's own intestinal flora, and then spread to cause urinary tract,intra-abdominal, and surgical wound infections. In severe cases,bacteremia may result with subsequent seeding of more distant sites.Whiteside, et al., Am. J. Infect. Control 11(4):125-9 (1983); Patterson,et al., Medicine (Baltimore) 74(4):191-200 (1995); Cooper, et al.,Infect. Dis. Clin. Practice 2:332-9. (1993). Recently in the UnitedStates, the National Nosocomial Infections Surveillance survey (NNIS)ranked Enterococci from the second to the fourth most common cause ofnosocomial infections. Enterococci frequently cause urinary tractinfections, bloodstream infections, and wound infections in hospitalizedpatients.

In addition, enterococci cause 5-15% of all bacterial endocarditiscases. Also, there is reported high prevalence of skin colonization withvancomycin-resistant enterococci that greatly increases the risk ofcatheter-related sepsis, cross-infection, or blood culturecontamination. CDC. National Nosocomial Infections Surveillance (NNIS)System report, Am. J. Infect. Control 26:522-33 (1998); Beezhold, etal., Clin. Infect. Dis. 24(4):704-6 (1997); Tokars, et al., Infect.Control Hosp. Epidemiol. 20(3):171-5 (1999). Of particular interest forthe NIMELS laser system are the infectious entities known as cutaneousor wound infections with Enterococci.

Enterococcal infections involve almost any skin surface on the bodyknown to cause skin conditions such as boils, carbuncles, bullousimpetigo and scalded skin syndrome. S. aureus is also the cause ofstaphylococcal food poisoning, enteritis, osteomilitis, toxic shocksyndrome, endocarditis, meningitis, pneumonia, cystitis, septicemia andpost-operative wound infections. Tomi, et al., J. Am. Acad. Dermatol.53(1):67-72 (2005); Breuer, et al., Br. J. Dermatol. 147(1):55-61(2002); Ridgeway, et al., J. Bone Joint Surg. Br. 87(6):844-50 (2005).Staphyloccoccus infections can be acquired while a patient is in ahospital or long-term care facility.

The confined population and the widespread use of antibiotics have ledto the development of antibiotic-resistant strains of S. aureus. Thesestrains are called methicillin resistant staphylococcus aureus (MRSA).Infections caused by MRSA are frequently resistant to a wide variety ofantibiotics and are associated with significantly higher rates ofmorbidity and mortality, higher costs, and longer hospital stays thaninfections caused by non-MRSA microorganisms. Risk factors for MRSAinfection in the hospital include surgery, prior antibiotic therapy,admission to intensive care, exposure to a MRSA-colonized patient orhealth care worker, being in the hospital more than 48 hours, and havingan indwelling catheter or other medical device that goes through theskin. Hidron, et al., Clin. Infect. Dis. 15;41(2):159-66 (2005); Hsueh,et al., Int. J. Antimicrob. Agents 26(1):43-49 (2005).

These enterococcal and staphylococcal infections have a huge potentialfor central venous catheters CVC Infection, and can cause substantialmorbidity and mortality in patients. Tomi, et al. (supra). In fact, thedata presents that in the United States, 15 million CVC days (i.e., thetotal number of days of exposure to CVCs by all patients in the selectedpopulation during the selected time period) occur in ICUs each yearMermel L A., Ann. Intern. 132:391-402 (2000). This translates into anaverage rate of CVC-associated bloodstream infections at 5.3 per 1,000catheter days in the ICU CDC (supra), or stated another way,approximately 80,000 CVC-associated bloodstream infections occur in ICUseach year in the United States. The attributable cost per infection tothe healthcare arena is an estimated $34,508-$56,000 Rello, et al., Am.J. Respir. Crit. Care Med. 162:1027-30 (2000); Dimick, et al., Arch.Surg. 136:229-34 (2001), and the annual cost of caring for patients withCVC-associated BSIs ranges from $296 million to $2.3 billion. Mermel LA., Ann. Intern. Med. 133:395 (2000).

The importance of fungal infections in the healthcare environment cannotbe overstated. As an example, Candida albicans is known to the seventhmost common pathogen associated with nosocomial infection in ICUpatients in hospitals. Fridkin, et al., Clinics In Chest Medicine,20:(2) (1999). With C. albicans the generally accepted therapeuticoptions for treatment are the polyene class of antifungals(amphotericin), the imidazole class of antifungals, and triazoles. Manyof these therapies need to be taken for extended periods of time (withconcurrent systemic and organ system danger) and there is much evidenceof emergence of antimicrobial-resistant fungal pathogens. When thisoccurs, the therapeutic options become few and limited.

As an example, there are patients with acquired immunodeficiencysyndrome patients, predominantly those with larger exposure to azoletherapy or low CD4 counts, that have developed azole-resistant C.albicans infections. Johnson, et al., J. Antimicrob. Chemother.35:103-114 (1995); Maenza, et al., J. Infect. Dis. 173:219-225 (1996).The recent appearance of azole-resistant C. albicans in acquiredimmunodeficiency syndrome patients most likely heralds coming resistanceissues in other immuno-compromised patient populations.

These data imply that the escalating use of prophylactic antifungaltherapy in highest risk patients for endogenous fungal infections maylead to the increasing frequency of fungal pathogens like C. krusei,which have intrinsic azole-resistance, or the even azole resistant C.glabrata or C. albicans. Maenza, et al., (supra); Beezhold, et al.,Clin. Infect. Dis. 24:704-706 (1997); Fridkin, et al., Clin. Microbiol.Rev. 9:499-511 (1996); Johnson, et al., J. Antimicrob. Chemother.35:103-114 (1995).

Continuing with this ominous trend, data from a 1998 multi-center studyof 50 U.S. medical centers, documents that 10% of C. albicans isolatesfrom the bloodstream of hospitalized patients were resistant to theantifungal drug fluconazole. Pfaller, et al., Diagn. Microbiol. Infect.Dis. 31:327-332 (1998). The resistant rate ranged from 5% to 15%,depending on the region of the United States, suggesting that localfactors, such as amount of azole usage, may play a role in the relativefrequency of azole-resistant C. albicans infections.

Of particular interest are the infectious entities known as cutaneousCandidiasis. These Candida infections involve the skin, and can occupyalmost any skin surface on the body. However, the most often occurrencesare in warm, moist, or creased areas (such as armpits and groins).Cutaneous candidiasis is extremely common. Huang, et al., Dermatol.Ther. 17(6):517-22 (2004). Candida is the most common cause of diaperrash, where it takes advantage of the warm moist conditions inside thediaper. The most common fungus to cause these infections is Candidaalbicans. Gallup, et al., J. Drugs Dermatol. 4(1):29-34 (2005). Candidainfection is also very common in individuals with diabetes and in theobese. Candida can also cause infections of the nail, referred to asonychomycosis, and infections around the corners of the mouth, calledangular cheilitis.

Thus, the literature described portends the need for innovative andnovel treatments to address these infections.

Traditionally, solid state diode lasers in the visible and near infraredspectrum (e.g., wavelengths of 600 nm to 1100 nm) have been used for avariety of purposes in medicine, dentistry, and veterinary sciencebecause of their preferential absorption curve for melanin andhemoglobin in biological systems. Because of the poor absorption inwater of near infrared optical energy, the penetration of such radiationin biological tissue is far greater than that of visible or longerinfrared wavelengths (e.g., mid-infrared and far infrared).Specifically, near infrared diode laser energy can penetrate biologicaltissue to about 4 centimeters. In contrast, longer wavelength radiantenergy (e.g., that of Er:YAG and CO₂ lasers producing mid infrared andfar infrared radiation, respectively), has a relatively high waterabsorption curve and penetrates biological tissue only to from 15 to 75microns (where 10,000 microns=1 cm). Thus, with radiation from nearinfrared diode lasers, heat deposition can occur much deeper inbiological tissue than for mid-infrared and far infrared wavelengths.Hence, it is more therapeutic for cancer treatment such aslaser-interstitial-thermal-therapy for deep tumor ablation orlaser-heat-generated-microbial sterilization.

For the destruction of bacterial cells with visible and near infrareddiode lasers, the prior art requires the presence of an exogenouschromophore at a site being irradiated and/or a very narrow therapeuticwindow and opportunity for treatment. Normal human temperature is 37°C., which corresponds to rapid bacterial growth in most bacterialinfections. When radiant energy is applied to a biological system with anear infrared diode laser, the temperature of the irradiated area startsto rise immediately, with each 10° C. rise carrying an injuriousbiological interaction. At 45° C. there is tissue hyperthermia, at 50°C. there is a reduction in enzyme activity and cell immobility, at 60°C. there is denaturation of proteins and collagen with beginningcoagulation, at 80° C. there is a permeabilization of cell membranes,and at 100° C. there is vaporization of water and biological matter. Inthe event of a significant duration of a temperature above 80° C., (5 to10 seconds in a local site), irreversible harm to healthy cells willresult.

Photothermolysis (heat induced lysis) of bacteria with near infraredlaser energy, in the prior art, requires a significant temperatureincrease that may endanger mammalian cells. However, most often it isdesired to destroy bacteria thermally, without causing irreversiblethermal damage to mammalian cells. Diode lasers have been used todestroy bacteria with visible laser energy (400 nm to 700 nm) in theprior art. The application to a bacterial site of exogenous chromophoreshas been needed for photodynamic therapy by visible radiation. In theprior art, photodynamic inactivation of bacteria has been achieved whenan exogenous chromophore is applied to prokaryotic (microbial) cells andis then irradiated with an appropriate light or laser source. Inreference to efforts to preferentially destroy bacteria by generation ofradical oxygen species with visible wavelengths coupled to an exogenouschromophore, two studies stand out in the prior art literature (see,e.g., Gibson et al., Clin. Infect. Dis., (16) Suppl 4:S411-3 (1993); andWilson et al., Oral Microb. Immunol. June; 8(3):182-7 (1993) and Wilsonet al., J. Oral. Pathol. Med. September; 22(8):354-7 (1993)).

Therefore, there is a need for improved modalities for the reduction ofmicrobial growth while minimizing damage to mammalian cells.

SUMMARY OF THE DISCLOSURE

The present invention provide methods, systems, and apparatus toselectively target a biological contaminant without intolerable risksand/or intolerable adverse effects on a biological moiety (e.g., amammalian tissue, cell or biochemical entity/preparations such as aprotein preparation) other than the biological contaminant.

The present invention provides method, systems, and apparatus that canapply near infrared radiant energy of certain wavelengths anddosimetries capable of impairing biological contaminants withoutintolerable risks and/or adverse effects to biological moieties otherthan a targeted biological contaminant associated with traditionalapproaches described in the art (e.g., loss of viability, orthermolysis). The methods, systems, and apparatus of the invention attimes are hereinafter referred by the acronym NIMELS (i.e., NearInfrared Microbial Elimination Laser System).

In a first aspect, the invention provides a method of reducing the levelof a biological contaminant in a target site without intolerable risksand/or intolerable adverse effects to biological moieties (e.g., amammalian tissue, cell or certain biochemical preparations such as aprotein preparation) in/at the given target site other than the targetedbiological contaminants, by irradiating the target site with opticalradiation of desired wavelength(s), power density level(s), and/orenergy density level(s). In certain embodiments, such applied opticalradiation may have a wavelength from about 850 nm to about 900 nm, at aNIMELS dosimetry, as described herein. In exemplary embodiments,wavelengths from about 865 nm to about 875 nm are utilized. In furtherembodiments, such applied radiation may have a wavelength from about 905nm to about 945 nm at a NIMELS dosimetry. In certain embodiments, suchapplied optical radiation may have a wavelength from about 925 nm toabout 935 nm. In representative non-limiting embodiments exemplifiedhereinafter, the wavelength employed is 930 nm. Biological contaminantsthat can be treated reduced and/or eliminated according to the presentinvention include microorganisms such as, for example, bacteria, fungi,molds, mycoplasmas, protozoa, prions, parasites, viruses, and viralpathogens. Exemplary embodiments, as noted below may employ multiplewavelength ranges including ranges bracketing 870 and 930 nm,respectively.

In a second aspect, the invention provides a method of reducing thelevel of a biological contaminant in a target site without intolerablerisks and/or intolerable adverse effects to biological moieties (e.g., amammalian tissue, cell or certain biochemical preparations such as aprotein preparation) located in/at the given target site other than thetargeted biological contaminants, by irradiating the target site with(a) an optical radiation having a wavelength from about 850 nm to about900 nm; and (b) an optical radiation having a wavelength from about 905nm to about 945 nm, at NIMELS dosimetries. With respect to thiscombination approach, and as discussed in more details hereinafter,embodiments of the invention can utilize wavelengths from about 865 nmto about 875 nm. Accordingly, in representative non-limiting embodimentsexemplified hereinafter, the wavelength employed is 870 nm. Similarly,with respect to the other wavelength ranges contemplated, the opticalradiation of the methods/systems described herein may utilize/produce awavelength from about 925 nm to about 935 nm and/or a second range,which as a non-limiting example could be about 865 nm to about 875 nm.In representative non-limiting embodiments exemplified hereinafter, thewavelength employed is 930 nm.

In the methods according to this aspect of the invention, irradiation bythe wavelength ranges contemplated may be performed independently, insequence, or essentially concurrently (all of which techniques canutilize pulsed and/or continuous-wave, CW, operation). Optical radiationcan be provided for a suitable NIMELS application time (Tn), e.g., offrom about 50 to about 450 seconds. Further, the NIMELS dosimetry mayadjusted as needed or according to formula or disometyr calculators. Forexample methods/systems according to the present disclosure may providesan energy density from about 200 J/cm² to about 700 J/cm² or an energydensity from about 275 J/cm² to about 500 J/cm².

In a third aspect, the invention provides a system to implement themethods according to other aspects of the invention, e.g., the first andthe second aspect of the invention. Such a system can include a laseroscillator for generating the radiation, a controller for calculatingand controlling the dosage of the radiation, and a delivery assembly(system) for transmitting the radiation to the treatment site through anapplication region. Suitable delivery assemblies/systems can includehollow waveguides, fiber optics, and/or free space/beam opticaltransmission components. Suitable free space/beam optical transmissioncomponents can include collimating lenses and/or aperture stops.

In one form, the system may utilize a two or more solid state diodelasers to function as a dual wavelength near-infrared optical source.The two or more diode lasers may be located in a single housing with aunified control, in exemplary embodiments. The two wavelengths caninclude emission in two ranges approximating 850 nm to 900 nm and 905 nmto 945 nm. The laser oscillator of the present invention may also beused to emit a single wavelength (or a peak value, e.g., centralwavelength) in either one of the ranges encompassed by the invention. Incertain embodiments, such a laser may be used to emit radiationsubstantially within the 865-875 nm and the 925-935 nm ranges asdescribed in more details with respect to the first and the secondaspects of the invention. Systems exemplified herein are provided asillustrations of possible embodiments of the invention, e.g., a systemdevised to emit radiation substantially at 870 nm and at 930 nm; otherwavelengths may be produced and utilized.

Systems according to the present invention can include a suitableoptical source for each individual wavelength range desired to beproduced. For a non-limiting example, a suitable solid stated laserdiode, a variable ultra-short pulse laser oscillator, or an ion-doped(e.g., with a suitable rare Earth element) optical fiber or fiber lasermay be used. In one form, a suitable near infrared laser can includetitanium-doped sapphire. Other suitable laser sources including thosewith other types of solid state, liquid, or gas gain (active) media maybe used within the scope of the present invention.

According to one embodiment of the present invention, a therapeuticsystem can include an optical radiation generation device adapted togenerate optical radiation substantially in a first wavelength rangefrom about 850 nm to about 900 nm, a delivery assembly for causing theoptical radiation to be transmitted through an application region, and acontroller operatively connected to the optical radiation generationdevice for controlling the dosage of the radiation transmitted throughthe application region, such that the time integral of the power densityand energy density of the transmitted radiation per unit area is below apredetermined threshold. Also contemplated according to this embodimentof the invention, are therapeutic systems especially adapted to generateoptical radiation substantially in a first wavelength range from about865 nm to about 875 nm.

According to further embodiments, a therapeutic system can include anoptical radiation generation device that is configured to generateoptical radiation substantially in a second wavelength range from about905 nm to about 945 nm; in certain embodiments the noted firstwavelength range may simultaneously or concurrently/sequentiallyproduced by the optical radiation generation device. Also contemplatedaccording to this embodiment are therapeutic systems especially adaptedto generate optical radiation substantially in a first wavelength rangefrom about 925 nm to about 935 nm.

The therapeutic system can further include a delivery assembly (system)for transmitting the optical radiation in the second wavelength range(and where applicable, the first wavelength range) through anapplication region, and a controller operatively for controlling theoptical radiation generation device to selectively generate radiationsubstantially in the first wavelength range or substantially in thesecond wavelength range or any combinations thereof.

According to a further embodiment, the controller of the therapeuticsystem includes a power limiter to control the dosage of the radiation.The controller may further include memory for storing patients' profileand dosimetry calculator for calculating the dosage needed for aparticular target site based on the information input by an operator. Inone embodiment, the memory may also be used to store information aboutdifferent types of diseases and the treatment profile, for example, thepattern of the radiation and the dosage of the radiation, associatedwith a particular application.

The optical radiation can be delivered from the therapeutic system tothe application site in different patterns. The radiation can beproduced and delivered as continuous wave (CW), or pulsed, or acombination of each. For example, in a single wavelength pattern or in amulti-wavelength (e.g., dual-wavelength) pattern. For a further example,two wavelengths of radiation can be multiplexed (optically combined) ortransmitted simultaneously to the same treatment site. Suitable opticalcombination techniques can be used, including, but not limited to, theuse of polarizing beam splitters (combiners), and/or overlapping offocused outputs from suitable mirrors and/or lenses, or other suitablemultiplexing/combining techniques. Alternatively, the radiation can bedelivered in an alternating pattern, in which the radiation in twowavelengths are alternatively delivered to the same treatment site. Aninterval between two or more pulses may be selected as desired accordingto NIMELS techniques of the invention. Each treatment may combine any ofthese modes of transmission. The intensity distributions of thedelivered optical radiation can be selected as desired. Exemplaryembodiments utilize top-hat or substantially top-hat (e.g., trapezoidal,etc.) intensity distributions. Other intensity distributions, such asGaussian may be used.

Other features and advantages of the present invention will be set forthin the detailed description of preferred embodiments that follow, and inpart will be apparent from the description or may be learned by practiceof the invention. Such features and advantages of the invention will berealized and attained by the systems, methods, and apparatusparticularly pointed out in the written description and claims hereof.

BRIEF DESCRIPTION OF THE DRAWINGS

For a fuller understanding of methods, systems, and apparatus of thepresent invention, reference is made to the following detaileddescription, which is to be taken with the accompanying drawings,wherein:

FIG. 1 is a double-logarithmic graph showing power density (ordinateaxis) versus irradiation time in seconds (abscissa axis). The mainlaser-tissue interactions are depicted as a function of different energydensity thresholds and parameters. The diagonal lines representdifferent energy densities showing energy density values exploitedaccording to the present invention (see circled area labeled NIMELS).

FIG. 2 illustrates a schematic diagram of a system according to oneembodiment of the present disclosure; and

FIGS. 3 a-3 d illustrate different patterns of optical radiationgenerated by the therapeutic system of the disclosure of FIG. 2.

FIG. 4 is a graphic representation of typical in vitro efficacy data (inpercent kill) obtained using representative methods, devices and systemsof the disclosure to target E. coli cells at different total energyvalues (in Joules).

FIG. 5 is a graphic representation of typical final sample temperatures(in ° C.) observed using representative methods and systems of thedisclosure to target E. coli cells at different total energy values (inJoules).

FIG. 6 is a graphic representation of typical final sample temperatures(in ° C.) observed in vitro using representative methods and systems ofthe disclosure to target S. aureus cells at different total energyvalues (in Joules).

FIG. 7 is a graphic representation showing typical in vitro efficacydata observed using representative methods and systems of the disclosureat thermally tolerable temperatures of the treated target site.

FIG. 8 is a diagram depicting the nail complex, showing the nail bed(matrix), the nail plate and the perionychium.

FIG. 9 is a diagram depicting the nail of a typical onychomycosispatient showing the plate, bed (sterile matrix and germinal matrix) andnail fold (lunula growing out under the eponychium) area beginning toimprove in the weeks following initial treatment according to one of theembodiments of the disclosure.

FIG. 10 is a diagram showing a chronically infected nail also showingcharacteristic features associated with chronic paronychia (e.g.,superficial infections in the epidermis bordering the nails).

FIG. 11 is a diagram depicting the nail of certain onychomycosispatients showing different discrete areas of the nail infected with apathogen, and other areas that are completely clean where the healthyportion of the nail plate is still hard and translucent.

FIGS. 12 a and c are schematic representation showing the illuminationpattern of a 1.5 cm irradiation spot with an incident Gaussian beampattern of the area of 1.77 cm². FIGS. 12 b and 12 d show by contrast,the uniform energy distribution (“Top-hat” pattern) used in certainembodiments of the disclosure, with the NIMELS laser system in vivo andin vitro.

FIG. 13 is a graph showing the Tn function for given spot-sizeparameters (1.2-2.2 cm diameter), treatment time parameters derived bydividing an energy density of 409 J/cm² by the power density, at a laseroutput power of 3.0 Watts.

FIG. 14 is a graph showing the Tn function for given spot-sizeparameters (1.2-2.2 cm diameter), treatment time parameters derived bydividing an energy density of 205 J/cm² by the power density, at a laseroutput power of 3.0 Watts.

FIG. 15 is a composite showing the improvement over time in theappearance of the nail of a typical onychomycosis patient treatedaccording to the methods of the disclosure.

FIG. 16 shows an embodiment of a NIMELS Optical Catheter Controllerincluding delivery assembly configured as multiple optical fibersembedded into the catheter controller around a catheter entry portplaced on a patient.

FIG. 17 shows a physical model constructed to simulate the embodiment ofFIG. 16.

FIG. 18 depicts the underside of a NIMELS Optical Catheter Controllersimilar to FIG. 16.

FIG. 19 shows a physical model according to FIG. 18, with the opticalfibers removed.

FIG. 20 is prototype enabled side view of a NIMELS Optical MicrobialCatheter Controller according to the present disclosure.

FIG. 21, is an additional view of the prototype of FIG. 20.

FIG. 22 is a further view of a NIMELS Optical Microbial CatheterController according to the present disclosure.

DETAILED DESCRIPTION OF THE DISCLOSURE

The patents, published applications, and scientific literature referredto herein establish the knowledge of those with skill in the art and arehereby incorporated by reference in their entirety to the same extent asif each was specifically and individually indicated to be incorporatedby reference. Any conflict between any reference cited herein and thespecific teachings of this specification shall be resolved in favor ofthe latter. Likewise, any conflict between an art-understood definitionof a word or phrase and a definition of the word or phrase asspecifically taught in this specification shall be resolved in favor ofthe latter.

As used in this specification, the singular forms “a”, “an” and “the”specifically also encompass the plural forms of the terms to which theyrefer, unless the content clearly dictates otherwise. For example,reference to “a NIMELS wavelength” includes any wavelength within theranges of the NIMELS wavelengths described, as well as combinations ofsuch wavelengths.

As used herein, unless specifically indicated otherwise, the word “or”is used in the “inclusive” sense of “and/or” and not the “exclusive”sense of “either/or.”

The term “about” is used herein to mean approximately, in the region of,roughly, or around. When the term “about” is used in conjunction with anumerical range, it modifies that range by extending the boundariesabove and below the numerical values set forth. In general, the term“about” is used herein to modify a numerical value above and below thestated value by a variance of 20%.

Technical and scientific terms used herein have the meaning commonlyunderstood by one of skill in the art to which the present disclosurepertains, unless otherwise defined. Reference is made herein to variousmethodologies and materials known to those of skill in the art. Standardreference works setting forth the general principles of microbiologyinclude, Joklik et al., Zinsser Microbiology, 20^(th) Ed., Appleton andLange (Prentice Hall), East Norwalk, Conn. (1992); Greenwood et al.,Medical Microbiology, 16^(th) Ed., Elsevier Science Ltd., New York(2003); Sambrook et al., Molecular Cloning: A Laboratory Manual, 2^(nd)Ed., Cold Spring Harbor Laboratory Press, New York, N.Y. (1989); Kaufmanet al., Eds., Handbook of Molecular and Cellular Methods in Biology inMedicine, CRC Press, Boca Raton, Fla. (1995); standard reference workssetting forth the general principles of pharmacology include Goodman andGilman's The Pharmacological Basis of Therapeutics, 10^(th) Ed., McGrawHill Companies Inc., New York, N.Y. (2001); and, standard dermatologyprinciples may be found in Habif et al., Skin Disease, Diagnosis andTreatment, 1^(st) Ed., Mosby, Inc., St. Louis, Mo. (2001); the entireteachings of all of which are incorporated herein by reference.

The present disclosure provides methods, systems, and apparatus to applynear infrared radiant energy of certain wavelengths and at a certaindosimetries as discussed herein capable of impairing targeted biologicalcontaminants with minimal risks to biological moieties other than thetargeted biological contaminant(s). Such methods and devices/systems forexample do not generate or rely on impermissible increases intemperatures (i.e., heat) associated with traditional approachesdescribed in the art.

Near infrared radiant energy has been used in the literature as opticaltweezers (Ashkin et al., Nature 330:769-771 (1987) the entire teachingsof which are incorporated herein by reference) used to manipulate andcontrol biological objects for a variety of applications for which itwas desirable to preserve the viability of the cells manipulated. Manyreported that the use of near infrared radiation as tweezers wasassociated with “opticution” or simply the undesired cell impairment (asmeasured for example by a quantifiable decrease in viability andproliferation) (Ashkin and Dziedzic, Ber. Bunsenges., Phys. Chem.93:254-260 (1989) the entire teachings of which are incorporated hereinby reference). In an effort of optimize optical tweezers that would nothamper the viability of the cells led to the discovery that the actionspectrum for photodamage exhibit maxima at 870 and 930 nm (Neuman etal., Biophys. J. 77:2856-2863 (1999) the entire teachings of which areincorporated herein by reference). Similar data in Chinese Hamster Ovary(“CHO”) cells (see, e.g., Liang et al., Biophys. J. 70:1529-1533 (1996))led investigators to believe that the wavelength dependence ofphotodamage seen in prokaryotic cells was shared by eukaryotic cells aswell (Neuman et al., Biophys. J. 77:2856-2863 (1999) the entireteachings of which are incorporated herein by reference). The consensusin the literature thus, has been that near infrared radiation havingwavelengths approximating or coinciding with identified maxima at 870and 900 nm causes cell damage in prokaryotic (e.g., bacteria) and ineukaryotic (e.g., CHO) cells.

More probing studies (exemplified hereinafter) using radiationapproximating and coinciding with the 870 and 900 maxima, has led to theelucidation of optical parameters (i.e., wavelength, power density,energy density, and duration of exposure) associated with a remarkabledifferential effect on targeted sites (e.g. cells). Using such dosimetryparameters it is now possible to use near infrared radiation to targetbiological contaminants while effecting other biological moieties onlymarginally, if at all. As it can be easily appreciated, such a discoveryhas many useful practical applications.

More specifically, it has been found that within certain dosimetryparameters, energy of a wavelength in the ranges of from about 905 nm toabout 945 nm is suitable to specifically target biological contaminantsin a target site without intolerable risks and/or intolerable adverseeffects to biological moieties in a given target site other than thetargeted biological contaminants.

Accordingly, in a first aspect, the disclosure provides a method ofreducing the level of a biological contaminant in a target site withoutintolerable risks and/or intolerable adverse effects to biologicalmoieties in a given target site other than the targeted biologicalcontaminants (e.g., a mammalian tissue, cell or certain biochemicalpreparations such as a protein preparation), by irradiating the targetsite with an optical radiation having a wavelength from about 905 nm toabout 945 nm. In certain embodiments the optical radiation may have awavelength from about 925 nm to about 935 nm. In representativenon-limiting embodiments exemplified hereinafter, the wavelengthemployed is 930 nm. The target site can include a medical device, whichmay be positioned in vivo, as described below in further detail.

It has also been found that the effects obtained by irradiating a targetsite with an optical radiation having a wavelength from about 905 nm toabout 945 nm may be augmented by also irradiating with at least oneadditional optical radiation with a wavelength from about 865 nm to 875nm at a NIMELS dosimetry. As evidenced herein, the combined irradiationfurther enhances the effect of the radiation in the 905-945 nm range byreducing the total energy and density required to obtain the desireddifferential effect on the treated target site. This finding isparticularly significant because it translates in a reduction of theradiation in the 905-930 nm range required to obtain the desired effect.As a result, this combined irradiation approach has the additionalbenefit of further minimizing intolerable risks and/or intolerableadverse effects to biological moieties other than the targetedbiological contaminants.

Such synergy has been found when target sites were subjected to twowavelengths of (a) from about 850 nm to 900 nm and of (b) from about 905nm to about 945 nm. In certain representative and non-limitingembodiments exemplified herein, it has been found that, at NIMELSdosimetries, irradiation with a wavelength in the 865-875 nm rangeenhances the effect of irradiation with a wavelength in the 925-935 nmrange. In certain embodiments, the target site was exposed to radiationswith λ=870 and λ=930 nm with a concomitant reduction of the requiredtotal energy and density.

NIMELS wavelengths as described above (e.g., 850-900 nm, and 905-945nm), may be used to irradiate the target site independently, insequence, and/or essentially concurrently.

Without wishing to be bound by any theory and not intending to limit anyaspect of the disclosure by any theory as to the underlying mechanismsresponsible for the phenomena observed, it is postulated that thewavelengths irradiated according to the present methods and systems areabsorbed by intracellular endogenous chromophores of prokaryotic andeukaryotic cells, and by the lipid bilayer in the cell membrane. It isfurther postulated that perhaps bacterial damage may be mediated viatoxic singlet oxygen and/or other reactive oxygen species.

It will be understood that intracellular endogenous chromophores ofprokaryotic and eukaryotic cells are lipid bilayers (plasma membranesand mitochondrial membranes) that contain large amounts of protein,cytochrome, and enzymatic inclusions, and that the lipid bilayers of thecontaminants and moieties targeted by the present invention will haveprotein/lipid rations of >1. Stated another way, none of the targetmembranes of the present invention in the contaminant or moiety containgreater than 49.99% lipid by dry weight.

As used herein, the expression “reducing the level of a biologicalcontaminant” is intended to mean a reduction in the level of at leastone active biological contaminant found in the target site being treatedaccording to the present disclosure. Empirically, a reduction of thelevel of a biological contaminant is quantifiably as a reduction of theviability of a biological contaminant in a target site (e.g., byhampering the viability of the subject biological contaminant and/or itsability to grow and/or divide). One of skills in the arts willappreciate that the expression “reduction of levels of a biologicalcontaminant” encompasses any reduction and need not be a 100% reduction.In certain embodiments in fact, the viability of a given biologicalcontaminant may only be reduced in part to allow other events to takeplace (e.g., allow a patient's immune system to react to a giveninfection, or allow other concomitant treatments—e.g., a systemicantibiotic treatment—to address a given infection). In certain instancesit has been found that a given biological contaminant's susceptibilityto antimicrobial may be enhanced following treatment according to thedisclosure. In particular embodiments, MRSA strains were found to bemore susceptible to antibiotics as a result of treatments according tothe disclosure.

As used herein, the term “biological contaminant” is intended to mean acontaminant that, upon direct or indirect contact with the target site,is capable of undesired and/or deleterious effect(s) on the target site(e.g., an infected tissue or organ of a patient) or upon a mammal inproximity of the target site (e.g., in the case of a cell, tissue, ororgan transplanted in a recipient, or in the case of a device used on apatient). Biological contaminants according to the disclosure aremicroorganisms such as, for example, bacteria, fungi, molds,mycoplasmas, protozoa, prions, parasites, viruses, and viral pathogensknown to those of skill in the art to generally be found in the targetsites according to the disclosure. One of skill in the art willappreciate that the methods and system/devices of the disclosure may beused in conjunction with a variety of biological contaminants known inthe literature at large (see, e.g., Joklik et al., (supra); andGreenwood et al., (supra)).

As used herein, the term “endogenous chromophores of biologicalcontaminants that are bacteria” is intended to mean cell membranes witha protein/lipid ratio of >1.

As used herein, the term “endogenous chromophores of biologicalcontaminants that are fungi” is intended to mean cell membranes andmitochondrial membranes with a protein/lipid ratio of >1.

The following lists are provided solely for the purpose of illustratingthe broad scope of microorganisms which may be targeted according to themethods and devices/systems of the disclosure and are not intended tolimit the scope of the applicability of the disclosure in any mannerwhatsoever.

Accordingly, illustrative non-limiting examples of biologicalcontaminants include any bacteria, such as, for example, Escherichia,Enterobacter, Bacillus, Campylobacter, Corynebacterium, Klebsiella,Treponema, Vibrio, Streptococcus and Staphylococcus.

To further illustrate, biological contaminants contemplated include anyfungus, such as, for example, Candida, Aspergillus, Cryptococcus,various dermatophytes (e.g., Trichophyton, Microsporum, andEpidermophyton), Coccidioides, Histoplasma, Blastomyces. Parasites mayalso be targeted biological contaminants such as Trypanosoma andmalarial parasites, including Plasmodium species, as well as molds;mycoplasmas; prions; and viruses, such as human immuno-deficiencyviruses and other retroviruses, herpes viruses, parvoviruses,filoviruses, circoviruses, paramyxoviruses, cytomegaloviruses, hepatitisviruses (including hepatitis B and hepatitis C), pox viruses, togaviruses, Epstein-Barr virus and parvoviruses.

It will be understood that the target site to be irradiated need not bealready infected with a biological contaminant. Indeed, the methods ofthe disclosure may be used “prophylactically,” prior to infection (e.g.,to prevent it). Exemplary embodiments may be used on medical devicessuch as catheters, artificial joints, etc.

In certain instances, irradiation may be palliative as well asprophylactic. Hence, the methods of the disclosure may be used toirradiate a tissue or tissues for a therapeutically effective amount oftime for treating or alleviating the symptoms of an infection. Theexpression “treating or alleviating” means reducing, preventing, and/orreversing the symptoms of the individual treated according to thedisclosure, as compared to the symptoms of an individual receiving nosuch treatment.

A practitioner will appreciate that the methods described herein are tobe used in concomitance with continuous clinical evaluations by askilled practitioner (physician or veterinarian) to determine subsequenttherapy. Hence, following treatment the practitioners will evaluate anyimprovement in the treatment of the underlying condition according tostandard methodologies. Such evaluation will aid and inform inevaluating whether to increase, reduce or continue a particulartreatment dose, mode of irradiation, and adjunctive treatments etc.

As discussed throughout the description of the disclosure, the term“target site” denotes any cell, tissue, organ, object or solution whichmay become contaminated with a biological contaminant. Thus, the targetsite may be a cell, tissue or organ of a mammal which is or may becomeinfected with a biological contaminant posing a risk to a mammal, e.g.,tissue surrounding an implanted (in vivo) medical device. In thealternative, the target site may be a cell, tissue or organ of a mammalwhich is or may become infected with a biological contaminant posing arisk to a mammal in proximity of the target site (e.g., such as forexample in the case of a cell, tissue, or organ transplanted in arecipient mammal, or in the case of a device used on a mammal). Foremostamong such mammals are humans, although the disclosure is not intendedto be so limited, and is applicable to veterinary uses. Thus, inaccordance with the disclosure, “mammals” or “mammal in need” or“patient” include humans as well as non-human mammals, particularlydomesticated animals including, without limitation, cats, dogs, andhorses. The target site may include a medical device, such as acatheter, stent, artificial joint, etc.

One of skill in the art will appreciate that the disclosure is useful inconjunction with a variety of diseases caused by or otherwise associatedwith any microbial, fungal, and viral infection (see in generalHarrison's, Principles of Internal Medicine, 13^(th) Ed., McGraw Hill,New York (1994) the entire teachings of which are incorporated herein byreference). In certain embodiments, the methods and the system accordingto the disclosure may be used in concomitance with traditionaltherapeutic approaches available in the art (see, e.g., Goodman andGilman's (supra)) to treat an infection by the administration of knownantimicrobial agents compositions. The terms “antimicrobialcomposition”, “antimicrobial agent” refer to the compounds andcombinations thereof that may be administered to an animal or human andwhich inhibit the proliferation of a microbial infection (e.g.,antibacterial, antifungal and antiviral).

The wide breath of applications contemplated include for example avariety of dermatological, podiatric, pediatric, and general medicine tomention but a few.

A plethora of dermatological conditions may be treated according to themethods, devices/systems of the disclosure (see, for example, Habif etal. (supra)). Without wishing to be bound to the specific infectionslisted, the disclosure for example may be used to treat Corynebacteriainfections which may cause erythrasma, trichomycosis axillaries, andpitted keratolysis; Staphylococcus infections which may cause impetigo,ecthyma and folliculitis, and Streptococcus infections that may causeimpetigo and erysipelas. Erythrasma is a superficial skin infectioncaused by Corynebacteria that commonly occurs in intertriginous spaces.Impetigo is a common infection in children that may also occur inadults. It is generally caused by either Staphylococcus aureus orStreptococcus. Ecthyma occurs in debilitated persons, such as patientswith poorly controlled diabetes, and is generally caused by the sameorganisms that cause impetigo. Patients with folliculitis present withyellowish pustules at the base of hairs, particularly on the scalp,back, legs and arms. Furuncles, or boils, are more aggressive forms offolliculitis. Erysipelas presents acutely as marked redness, pain andswelling in the affected area. The illness is generally believed to becaused by beta-hemolytic Streptococci. See, for example, Trueb et al.,Pediatr Dermatol 1994;11:35-8 (1994); Trubo et al., Patient Care31(6):78-94 (1997); Chartier et al., Int. J. Dermatol. 35:779-81 (1996);and Eriksson et al., Clin. Infect. Dis. 23:1091-8 (1996) the entireteachings of which are incorporated herein by reference.

Similarly, fungus and yeast may infect skin tissues causing a variety ofconditions (dermatomycoses) which may be addressed according to thedisclosure including, for example, tinea capitis, tinea barbae, tineacruris, tinea manus, tinea pedis and tinea unguium (see, onychomycosisdiscussed infra) (see, Ansari et al., Lower Extremity Wounds 4(2):74-87(2005); Zaias, et al., J. Fam. Pract. 42:513-8 (1996), Drake et al., J.Am. Acad. Dermatol. 34(2 Pt 1):282-6 (1996); Graham et al., J. Am. Acad.Dermatol. 34(2 Pt 1):287-9 (1996); Egawa et al., Skin Research and Tech.12:126-132 (2005); and Hay, Dermatol. Clin. 14:113-24 (1996) the entireteachings of which are incorporated herein by reference). Candidalpathogen based infections will generally occur in moist areas, such as,skinfolds and diaper area. Cutaneous wounds that are caused by woodsplinters or thorns may result in sporotrichosis (see, Kovacs et al.,Postgrad Med 98(6):61-2,68-9,73-5 (1995) the entire teachings of whichare incorporated herein by reference). Candida albicans andTrichophyton, Epidermophyton, Microsporum, Aspargillum, and Malasseziaspecies are the common infecting organisms (see, Masri-Fridling,Dermatol. Clin. 14:33-40 (1996) the entire teachings of which areincorporated herein by reference).

HPV (Human papillomavirus) may also cause skin infections that maymanifest clinically as different types of warts, depending on thesurface infected and its comparative moisture. Commonly occurring wartsinclude common warts, plantar warts, juvenile warts and condylomata. Nostandard and routinely effective treatment for warts exists to date(Sterling, Practitioner 239:44-7 (1995) the entire teachings of whichare incorporated herein by reference).

As exemplified hereinafter, the disclosure may be used for the treatmentof onychomycosis i.e., a disease (e.g., a fungal infection) of the nailplate on the hands or feet. As used herein, reference to a “nail”includes reference to one, or some, or all parts of the nail complex,including the nail plate (the stratum corneum unguis, which is the hornycompact outer layer of the nail, i.e., visible part of the nail), thenail bed (the modified area of the epidermis beneath the nail plate,over which the nail plate slides as it grows), the cuticle (the tissuethat overlaps the nail plate and rims the base of the nail), the nailfolds (the skin folds that frame and support the nail on three sides),the lunula (the whitish half-moon at the base of the nail), the matrix(the hidden part of the nail under the cuticle), and the hyponychium(the thickened epidermis underneath the free distal end of a nail) andthe nail matrix. Nails grow from the matrix. Nails are composed largelyof keratin, a hardened protein (that is also in skin and hair). As newcells grow in the matrix, the older cells are pushed out, compacted andtake on the familiar flattened, hardened form of a fingernail ortoenail.

Nail fungal disease may be caused by the three genera of dermatophytes,Trichophyton, Microsporum, Epidermophyton, the yeast Candida, (the mostprevalent of which being C. albicans, and/or or moulds such asScopulariopsis brevicaulis, Fusarium spp., Aspergillus spp., Alternaria,Acremonium, Scytalidinum dimidiatum (Hendersonula toruloides),Scytalidinium hyalinum. Onychomycosis may affect one or more toenailsand/or fingernails and most often involves the great toenail or thelittle toenail. It can present in one or several different patterns suchas lateral onychomycosis (a white or yellow opaque streak appears at oneside of the nail), subungual hyperkeratosis (scaling occurs under thenail), and distal onycholysis (when the end of the nail lifts upwards).Common clinical findings include crumbling of the free edge (e.g.,superficial white onychomycosis), flaky white patches and pits appear onthe top of the nail plate (e.g., proximal onychomycosis), yellow spotsappear in the half-moon (lunula), and the complete destruction of thenail (see Sehgal and Jain, Inter. J. of Dermatol. 39:241-249 (2000);Hay, JEADV 19 (Suppl. 1.):1-7 (2005); Warshaw et al., Inter. J. ofDermatol. 44:785-788 (2005); Sigureirsson et al., J. of Dermatol.Treatmt. 17:38-44 (2006); Rodgers et al., Amer. Fam. Phys.; see, athttp://www.aafp.org/afp/20010215/663.html)); Lateur, J. of Cosmet.Dermatol. 5:171-177 (2006) the entire teachings of which areincorporated herein by reference).

It will be readily appreciated that treatment according to thedisclosure also provides modalities to address many known clinicalevents associated with onychomycosis and tinea corporis. The absence ofeffective therapy for many patients affected by onychomycosis has beenfound to have a profound impact on the patients' quality of life leadingto considerable psychological and psychosocial consequences (see, e.g.,Elewski et al., Int. J. Dermatol. 36:754-756 (1997) the entire teachingsof which are incorporated herein by reference). Treatment according tothe instant disclosure thus, provide a much needed relief from theliterature-recognized impact these diseases have on self-image andoverall life quality.

Reports in the literature have also confirmed that fungal infections(e.g., onychomycosis) is a risk factor for bacterial tissue infectionsincluding infections such as for example acute bacterial cellulitis(see, e.g., Roujeau et al., Dermatology 209:301-307 (2004) the entireteachings of which are incorporated herein by reference). Treatment offungal infections as described herein therefore provides a novelapproach to curb secondary or concomitant infections.

It has been recognized that the significance of onychomycosis and tineacorporis in the diabetic patient may lead to infections, especiallybacterial sepsis which may turn into a life-threatening problem giventhe susceptibility and propensity of diabetic patients to secondaryinfections at large (see, e.g., Rich, J. Am. Acad. Dermatol. 35:S10-12(1996) the entire teachings of which are incorporated herein byreference). In patients with labile diabetes, recurrent candidiasis canresult in candida sepsis and ultimately may also lead to candidaparonchia further complicating the nail dystrophy from long standingonychomycosis (see, e.g., Millikan et al., Int. J. Dermatol. 38(2):13-16(1999) the entire teachings of which are incorporated herein byreference).

Numerous nails that are chronically infected with a pathogen often alsosuffer from chronic or acute paronychia (see, e.g., Rockwell, AmericanMed. Physic. 63(6):1113-1116 (2001); and Grover et al., Dermatol. Surg.32393-399 (2006) the entire teachings of which are incorporated hereinby reference). Chronic paronychias are localized, superficial infectionsof the perionychium (epidermis bordering the nails). Paronychialinfections develop when a disruption occurs between the seal of theproximal nail fold and the nail plate that allows a portal of entry forinvading organisms. Chronic paronychia is generally nonsuppurative andis a difficult disease to treat. Chronic paronychia as a rule, causesswollen, red, tender and boggy nail folds where the symptoms of thedisease present for six weeks or longer and are concominent with longterm onychomycosis. The disease causing pathogen in these casestypically is a Candida species.

In accordance with some embodiments, the methods and devices/systems ofthe disclosure may be used in conjunction with the administration of apharmaceutically active compound and/or a composition containing apharmaceutically active compound. Such administration may be systemic ortopical. Various such antifungal pharmaceutically active compounds andcompositions suitable for systemic (e.g., orally or by parenteraladministration) or topical (e.g., ointments, creams, sprays, gels,lotions and pastes) are known in the art. See, for example, terbinafineas described in e.g., U.S. Pat. Nos. 4,755,534; 6,121,314; 4,680,291;5,681,849; 5,856,355; 6,005,001, and itraconazole as described in e.g.,U.S. Pat. Nos. 5,633,015; 4,727,064; 5,707,975; the entire teachings ofwhich are incorporated herein by reference.

As illustrated infra, it has been found that antibiotic resistantbacteria may be effectively treated according to the methods of thedisclosure. In addition, it has been found that the methods of thedisclosure may be used to augment traditional approaches to be used incombination with, in lieu of, or even serially as effective therapeuticapproaches. Accordingly, the disclosure may be combined with antibiotictreatment. The term “antibiotic” includes, but is not limited to,β-lactams penicillins and cephalosporins), vancomycins, bacitracins,macrolides (erythromycins), ketolides (telithromycin), lincosamides(clindomycin), chloramphenicols, tetracyclines, aminoglycosides(gentamicins), amphotericns, cefazolins, clindamycins, mupirocins,sulfonamides and trimethoprim, rifampicins, metronidazoles, quinolones,novobiocins, polymixins, oxazolidinone class (e.g., linezolid),glycylcyclines (e.g., tigecycline), cyclic lipopeptides (e.g.,daptomycin), pleuromutilins (e.g., retapamulin) and gramicidins and thelike and any salts or variants thereof. It also understood that it iswithin the scope of the present disclosure that the tetracyclinesinclude, but are not limited to, immunocycline, chlortetracycline,oxytetracycline, demeclocycline, methacycline, doxycycline andminocycline and the like. It is also further understood that it iswithin the scope of the present disclosure that aminoglycosideantibiotics include, but are not limited to, gentamicin, amikacin andneomycin and the like.

Other known approaches to the treatment of microbial infectionscontemplated in conjunction with the methods, devices, and systemsdescribed herein include the use of suitable medical dressings. The term“medical dressing” as used herein refers to any covering, protective orsupportive, for diseased or injured parts of the skin, or internalorgans of a human or animal. The dressing can be, but is not limited to,an absorbent dressing such as a gauze, a sterilized gauze or absorbentcotton, an antiseptic dressing permeated with an antiseptic solution todelay or prevent the onset of an infection, a dry dressing comprising adry gauze, dry absorbent cotton or any other dry material that may besterilized by any means known to one of ordinary skill in the art andwhich does not render the dressing unacceptable for placing over an openwound. The medical dressing as understood by the present disclosure mayalso comprise a non-adherent dressing that will not adhere to aninfected wound or injury, a protective dressing intended to preventfurther injury or infection to the infected part of the body, and a wetdressing wherein the dressing is wetted before application to theinfected site. The term “medical dressing” may further include anoil-based support such as vitamin E in which an antimicrobialcomposition according to the present disclosure is dissolved. Theoil-base such as, for example, vitamin E can form a barrier to furthermicrobial infection and will leach an antimicrobial composition into thedamaged tissue.

In certain instances, the methods, devices, and systems of thedisclosure may be used to disinfect/sterilize or maintain a givenproduct essentially ‘microbe-free’. Accordingly, a target site may alsobe an object such as, for example, a medical device (e.g., a catheter ora stent), an artificial prosthetic device (e.g., an artificial joint).

Biofilms on indwelling medical devices can contain populations ofgram-positive or gram-negative bacteria or fungi. Gram positiveorganisms encountered in medical device biofilms are E. faecalis, S.aureus, S. epidermidis, and S. viridans. Gram-negative bacteriaencountered are E. coli, K. pneumoniae, Proteus mirabilis, and P.aeruginosa. These bacteria can are generally derived from the skin ofpatients or healthcare workers, tap water to which entry ports areexposed, or other sources in the environment such as the patients ownstool.

Bacterial biofilms grow when microorganisms irreversibly adhere to a wetsurface (such as the internal lumen of a catheter) and produceextracellular polymers that assist adhesion and provide a structuralmatrix for the colony. The surface that biofilms form on may be inert,nonliving material or living tissue. Microorganisms in a biofilm, behavedifferently from planktonic (freely suspended) bacteria regarding growthrates and ability to resist antimicrobial treatments, and consequentlypose a major medical and public health problem. The present disclosurecan inhibit planktonic bacteria from attaching to the surface of amedical device and hence prevent formation of a microbial biofilm.

There are a number of variables that aid in the establishment of whethera contaminated indwelling medical device will develop a biofilm. Onefactor is that the bacteria or fungus adhere to the exposed surfaces ofthe device if the exposure time is long enough, and consequently thebacteria or fungus become irreversibly attached. As an example of theproblem, urinary catheters (tubular latex or silicone devices), wheninserted readily obtain biofilms on the inner or outer surfaces of thecatheter. The organisms commonly contaminating these devices anddeveloping biofilms are S. epidermidis, E. faecalis, E. coli, P.mirabilis, P. aeruginosa, K. pneumoniae, and other gram-negativeorganisms. The longer the urinary catheter remains in place, the greaterthe tendency of these organisms to develop biofilms and result inurinary tract infections, a large medical problem.

The prior art has suggested a number of ways to prevent the occurrenceof biofilms in catheters. The conventional methods include usingmeticulous aseptic technique during implantation, topical antibiotics atthe insertion site, minimizing the duration of catheterization, makinguse of an in-line filter for intravenous fluids, creating mechanicalbarriers to prevent influx of organisms by attaching the catheter to asurgically implanted cuff, and attempting to coating the inner lumen ofthe catheter with an antimicrobial agent. However, none of the prior artmethods works as effectively as desired.

The methods, systems, and apparatus according to the present disclosurethus, can be used with in-dwelling medical devices such as for examplecentral venous catheters and needleless connectors, endotracheal tubes,peritoneal dialysis catheters, tympanostomy tubes, urinary catheters,and stents, tec. to prevent/mitigate biofilm formation or reduce otherbiological contaminants as described herein for such devices. Other suchmedical devices can include, but are not limited to, an IV catheter, acentral venous line, an arterial catheter, a peripheral catheter, adialysis catheter, an external fixator pin, peritoneal dialysiscatheter, an epidural catheter, a chest tube, and/or a gastronomyfeeding tube

Embodiments of the disclosure may also be used to treat biochemical orchemical materials which are infected or may become infected with abiological contaminant (e.g., biochemical or pharmaceutical solution).Most of the methods in the art used to produce preparations to be usedin mammals (e.g., immunoglobulin preparations) may result incontamination of the product by pathogens (i.e., biologicalcontaminants). For example, monoclonal immunoglobulin preparations aremade in one of three general fashions. The first involves production ina cell culture system, the second uses an animal as a temporarybioreactor for monoclonal immunoglobulin production, and the thirdinvolves inserting the gene for a desired monoclonal immunoglobulin intoan animal in such a manner as to induce continuous production of themonoclonal immunoglobulin into a fluid or tissue of the animal so thatit can be continuously harvested (transgenic production). In the contextof the first method, the cells producing the monoclonal immunoglobulinmay harbor undetected viruses that can be produced in the culturesystem. Both of the remaining methods involve the use of an animal toeither serve as a host for the monoclonal immunoglobulin-producing cellsor as a bioreactor to manufacture the monoclonal immunoglobulin productitself. Obviously, these products face the risk of contamination bypathogens infecting or harbored by the host animal. Such pathogensinclude, viruses, bacteria, yeasts, molds, mycoplasmas, and parasites,among others. Consequently, it is of importance that any biologicallyactive contaminant in the monoclonal immunoglobulin product beinactivated before the product is used. This is especially importantwhen the product is to be administered directly to a patient. This isalso critical for various monoclonal immunoglobulin products which areprepared in media which contain various types of plasma and which may besubject to mycoplasma or other viral contaminants.

Among the viruses of concern for both human and animal-derivedbiologics, the smallest viruses of concern belong to the family ofParvoviruses and the slightly larger protein-coated Hepatitis virus. Inhumans, the Parvovirus B19, and Hepatitis A, as well as larger and lesshardy viruses such as HIV, CMV, Hepatitis B and C and others, are theagents of concern. In porcine-derived products and tissues, the smallestcorresponding virus is Porcine Parvovirus.

The interaction between the target site being treated and the energyimparted is defined by a number of parameters including: thewavelength(s); the chemical and physical properties of the target site;the power density or irradiance of beam; whether a continuous wave (CW)or pulsed irradiation is being used; the laser beam spot size; theexposure time, energy density, and any change in the physical propertiesof the target site as a result of laser irradiation with any of theseparameters. In addition, the physical properties (e.g., absorption andscattering coefficients, scattering anisotropy, thermal conductivity,heat capacity, and mechanical strength) of the target site may alsoaffect the overall effects and outcomes.

The term “NIMELS dosimetery” denotes the power density (W/cm²) and theenergy density (J/cm²) (where 1 Watt=1 Joule per second) values at whicha subject wavelength according to the disclosure is capable of reducingthe level of a biological contaminant in a target site withoutintolerable risks and/or intolerable side effects on a biological moiety(e.g., a mammalian cell, tissue, or organ) other than the biologicalcontaminant.

As show in FIG. 1 (reproduced in part from Boulnois, Lasers Med. Sci.1:47-66 (1986) the entire teachings of which are incorporated herein byreference), at low power densities (also referred to as irradiances)and/or energies, the laser-tissue interactions can be described aspurely optical (photochemical), whereas at higher power densitiesphoto-thermal interactions ensue. In certain embodiments exemplifiedhereinafter, NIMELS dosimetry parameters lie between known photochemicaland photo-thermal parameters (see, FIG. 1), in an area traditionallyused for photodynamic therapy in conjunction with exogenous drugs, dyesat large, and/or chromophores.

As shown in FIG. 1 depending on the interaction, the energy density—alsoexpressible as fluence, or the product (or integral) of particle orradiation flux and time—for medical laser applications in the arttypically varies between 1 J/cm² and 10,000 J/cm² (five orders ofmagnitude), whereas the power density (irradiance) varies from 1×10⁻³W/cm² to over 10¹² W/cm² (15 orders of magnitude). Upon taking thereciprocal correlation between the power density and the irradiationexposure time, it can be observed that approximately the same energydensity is required for any intended specific laser-tissue interaction.As a result, laser exposure duration (irradiation time) is the primaryparameter that determines the nature and safety of laser-tissueinteractions. For example, if one were mathematically looking for athermal vaporization of tissue in vivo (non-ablative) as thelaser-tissue interaction of choice for a particular therapy, (based onBoulnois 1986), it can be seen that to produce an energy density of 1000J/cm² (within the thermal-vaporization shaded area of FIG. 1) one coulduse any of the following dosimetry parameters:

TABLE I Example of Values Derived on the Basis of the Boulnois TablePOWER DENSITY TIME ENERGY DENSITY 1 × 10⁵ W/cm² 0.01 sec. 1000 J/cm² 1 ×10⁴ W/cm² 0.10 sec. 1000 J/cm² 1 × 10³ W/cm² 1.00 sec. 1000 J/cm²

This progression describes a suitable method/technique or basicalgorithm to be used for a NIMELS interaction against a biologicalcontaminant in a tissue. In other words, this mathematical relation is areciprocal correlation to achieve a laser-tissue interaction phenomena.This logic is used as a basis for dosimetry calculations for theobserved (through experimentation) antimicrobial phenomenon imparted byNIMELS energies with insertion of NIMELS experimental data in the energydensity and time and power parameters.

On the basis of the particular interactions at the target site beingirradiated (such as the chemical and physical properties of the targetsite; whether continuous wave (CW) or pulsed irradiation is being used;the laser beam spot size; and any change in the physical properties ofthe target site—e.g., absorption and scattering coefficients, scatteringanisotropy, thermal conductivity, heat capacity, and mechanicalstrength—, as a result of laser irradiation with any of theseparameters), a practitioner is able to adjust the power density and timeto obtain the desired energy density.

The examples provided herein show such relationships in the context ofboth in vitro and in vivo treatments. Hence, in the context of thetreatment of onychomycosis, for spot sizes having a diameter of 1-4 cm,power density values were varied from about 0.5 W/cm² and 5 W/cm² tostay within safe and non-damaging/minimally damaging thermallaser-tissue interactions well below the level of “denaturization” and“tissue overheating”. Other suitable spot sizes may be used.

With this reciprocal correlation, the threshold energy density neededfor a NIMELS interaction with these wavelengths can be maintainedindependent of the spot-size so long as the desired energies aredelivered. In exemplary embodiments, the optical energy is deliveredthrough a uniform geometric distribution to the tissues (e.g., aflat-top, or top-hat progression). With such a technique (logic) inmind, a suitable NIMELS dosimetry sufficient to generate a NIMELS effectcan calculated to reach the threshold energy densities required toreduce the level of a biological contaminant but below the level of“denaturization” and “tissue overheating”.

NIMELS Dosimetries exemplified herein to target microbes in vivo, were200 J/cm²-700 J/cm² for approximately 100 to 700 seconds. These powervalues do not approach power values associated with photoablative orphotothermal (laser/tissue) interactions.

The intensity distribution of a collimated laser beam is given by thepower density of the beam, and is defined as the ratio of laser outputpower to the area of the circle in (cm²). As illustrated in FIGS. 12Aand 12C, the illumination pattern of a 1.5 cm irradiation spot with anincident Gaussian beam pattern of the area of 1.77 cm² may produce atleast six different power density values within the 1.77 cm² irradiationarea.

These varying power densities increase in intensity (or concentration ofpower) over the surface area of the spot from 1 (on the outer periphery)to 6 at the center point. In certain embodiments of the disclosure, abeam pattern is provided which overcomes this inherent error associatedwith traditional laser beam emissions. FIGS. 12B and 12D show a uniformenergy distribution (the “top-hat” pattern as mentioned infra) used incertain embodiments of the disclosure to obtain more consistent powerenergy values in the irradiation area.

As shown in FIGS. 12B and 12D, in exemplary embodiments, a NIMELS lasersystem can correct for this error by illuminating in a uniform pattern(top-hat, or a 2π angular step intensity distribution) over an extendedarea, to insure that there are no or minimal “hot-spots” or “cold spots”in the three dimensional distribution pattern of energy that couldnegatively interfere with treatment by burning the tissue in the middleof the spot or having a sub-therapeutic energy density on the periphery.Other embodiments may utilize substantially top-hat, e.g., trapezoidal,Gaussian, or other suitable intensity distributions.

In the alternative, NIMELS parameters may be calculated as a function oftreatment time (Tn) as follows: Tn=Energy Density/Power Density.

In certain (see e.g., the in vitro experiments exemplified herein)embodiments Tn is of from about 50 to about 300 seconds; in otherembodiments, Tn is from about 75 to about 200 seconds; in yet otherembodiments, Tn is from about 100 to about 150 seconds. In other in vivoembodiments Tn is from about 100 to about 450 seconds.

Utilizing the above relationships and desired optical intensitydistributions, e.g., flat-top illumination geometries as describedherein, a series of in vivo energy parameters has been experimentallyproven as effective for NIMELS microbial decontamination therapy invivo. These are shown below for a fixed laser output power of 3 Watts oflaser energy for a NIMELS treatment. A key parameter for a given targetsite has thus been shown to be the energy density required for NIMELStherapy at a variety of different spot sizes and power densities.

Hence, “NIMELS dosimetery” encompasses ranges of power density and/orenergy density from a first threshold point at which a subjectwavelength according to the disclosure is capable of optically reducingthe level of a biological contaminant in a target site to a secondend-point immediately before those values at which an intolerableadverse risk or effect is detected (e.g., thermal damage such as, forexample, poration) on a biological moiety. One of skill in the art willappreciate that under certain circumstances certain adverse effectsand/or risks on a target site (e.g., a mammalian cell, tissues, ororgan) may be tolerated in view of the inherent benefits accruing fromthe methods of the disclosure. Accordingly, the end point contemplatedare those at which the adverse effects are considerable and thus,undesired (e.g., cell death, protein denaturation, DNA damage,morbidity, or mortality).

In certain embodiments, e.g., for in vivo applications, the powerdensity range contemplated herein is from about 0.25 to about 40 W/cm².In other embodiments, the power density range is from about 0.5 W/cm² toabout 25 W/cm².

In further embodiments, power density ranges can encompass values fromabout 0.5 W/cm² to about 10 W/cm². Power densities exemplified hereinare from about 0.5 W/cm² to about 5 W/cm². Power densities in vivo from1.5-2.5 W/cm² have been shown to be effective for various bacteria.

Empirical data appears to indicate that higher power density values aregenerally used when targeting a biological contaminant in an in vitrosetting (e.g., plates) rather than in vivo (e.g., toe nail).

In certain embodiments (see in vitro examples), the energy density rangecontemplated herein is greater than 50 J/cm² but less than about 25,000J/cm². In other embodiments, the energy density range is from about 750J/cm² to about 7,000 J/cm². In yet other embodiments, the energy densityrange is from about 1,500 J/cm² to about 6,000 J/cm² depending onwhether the biological contaminant is to be targeted in an in vitrosetting (e.g., plates) or in vivo (e.g., toe nail or surrounding amedical device).

In certain embodiments (see in vivo examples), the energy density isfrom about 100 J/cm² to about 500 J/cm². In yet other in vivoembodiments, the energy density is from about 175 J/cm² to about 300J/cm². In yet other embodiments, the energy density is from about energydensity from about 200 J/cm² to about 250 J/cm². In some embodiments,the energy density is from about 300 J/cm² to about 700 J/cm². In someother embodiments, the energy density is from about 300 J/cm² to about500 J/cm². In yet others, the energy density is from about 300 J/cm² toabout 450 J/cm².

Power densities empirically tested for various in vitro treatment ofmicrobial species were from about 1 W/cm²to about 20 W/cm².

One of skill in the art will appreciate that the identification ofparticularly suitable NIMELS dosimetry values within the power densityand energy density ranges contemplated herein for a given circumstancemay be empirically done via routine experimentation and even by meretrial and error as it is currently done in several presently-availablelaser uses. Practitioners (e.g., dentists) using near infrared energiesin conjunction with periodontal treatment routinely adjust power densityand energy density based on the exigencies associated with each givenpatient (e.g., adjust the parameters as a function of tissue color,tissue architecture, and depth of pathogen invasion). As an example,laser treatment of a periodontal infection in a light-colored tissue(e.g., a melanine deficient patient) will have greater thermal safetyparameters than darker tissue, because the darker tissue will absorbnear-infrared energy more efficiently, and hence transform thesenear-infrared energies to heat in the tissues faster. Hence the obviousneed for the ability of a practitioner to identify multiple differentNIMELS dosimetry values for different therapy protocols.

Any suitable materials (e.g., laser active media, resonatorconfiguration, etc.) and/or methods known to those of skill can beutilized in carrying out the present disclosure. Certain exemplarymaterials, methods, and configurations are described. Materials,reagents and the like to which reference is made in the followingdescription and examples are obtainable from commercial sources, unlessotherwise noted.

In a further aspect, the present disclosure provides a therapeuticradiation system (i.e., the NIMELS system). FIG. 2 illustrates aschematic diagram of a therapeutic radiation treatment device accordingone embodiment of the present disclosure. The therapeutic system 10includes an optical radiation generation device 12, a delivery assembly14, an application assembly (or region) 16, and a controller 18.According one aspect of the present disclosure, the optical radiationgeneration device (source) includes one or more suitable lasers, L1 andL2. A suitable laser may be selected based on a degree of coherence

In exemplary embodiments, a therapeutic system can include at least onediode laser configured and arranged to produce an output in the nearinfrared region. Suitable diode lasers can include a semiconductormaterials selected from among In_(x)Ga_(1-x)As, GaAs_(1-x)P_(x),Al_(x)Ga_(1-x)As, and (Al_(x)Ga_(1-x))_(y)In_(1-y)As, for producingradiation in desired wavelength ranges, e.g., 850 nm-900 nm and 905nm-945 nm (where within each semiconductor alloy, x and y indicatefractions of 1). Suitable diode laser configurations can includecleave-coupled, distributed feedback, distributed Bragg reflector,vertical cavity surface emitting lasers (VCSELS), etc.

With continued reference to FIG. 2, in certain embodiments the deliveryassembly 14 can generate a “flat-top” energy profiles for uniformdistribution of energy over large areas. As noted, the optical radiationgeneration device 12 can include one or more lasers, e.g., laseroscillators L1 and L2. In exemplary embodiments, one laser oscillatorcan be configured to emit optical radiation in a first wavelength rangeof 850 nm to 900 nm, and the other laser oscillator can be configured toemit radiation in a second wavelength range of 905 nm to 945 nm. Incertain embodiments, one laser oscillator is configured to emitradiation in a first wavelength range of 865 nm to 875 nm, and the otherlaser oscillator 28 is configured to emit radiation in a secondwavelength range of 925 nm to 935 nm. The geometry or configuration ofthe individual laser oscillators may be selected as desired, and theselection may be based on the intensity distributions produced by aparticular oscillator geometry/configuration.

With continued reference to FIG. 2, in certain embodiments, the deliveryassembly 14 includes an elongated flexible optical fiber adapted fordelivery of the dual wavelength radiation from the oscillators 26 and 28to the application region 16. See also, FIGS. 16 and 17. The deliveryassembly 14 may have different formats (e.g., including safety featuresto prevent thermal damage) based on the application requirements. Forexample, in one form, the delivery assembly 14 may be constructed with aminimized size and with a shape for inserting into a patient's body.

In alternate forms, the delivery assembly 14 may be constructed with aconical shape for emitting radiation in a diverging-conical manner toapply the radiation to a relatively large area. Hollow waveguides may beused for the delivery assembly 14 in certain embodiments. Other size andshapes of the delivery assembly 14 may also be employed based on therequirements of the application site. In exemplary embodiments, thedelivery assembly 14 can be configured for free space or free beamapplication of the optical radiation, e.g., making use of availabletransmission through tissue at NIMELS wavelengths described herein. Forexample, at 930 nm (and to a similar degree, 870 nm), the appliedoptical radiation can penetrate patient tissue by up to 1 cm or more.Such embodiments may be particularly well suited for use with in vivomedical devices as described below.

In an exemplary embodiment, the controller 18 includes a power limiter24 connected to the laser oscillators L1 and L2 for controlling thedosage of the radiation transmitted through the applicationassembly/region 16, such that the time integral of the power density ofthe transmitted radiation per unit area is below a predeterminedthreshold, which is set up to prevent damages to the healthy tissue atthe application site. The controller 18 may further include a memory 26for storing treatment information of patients. The stored information ofa particular patient may include, but not limited to, dosage ofradiation, (for example, including which wavelength, power density,treatment time, skin pigmentation parameters, etc.) and application siteinformation (for example, including type of treatment site (lesion,cancer, etc.), size, depth, etc.).

In an exemplary embodiment, the memory 26 may also be used to storeinformation of different types of diseases and the treatment profile,for example, the pattern of the radiation and the dosage of theradiation, associated with a particular type of disease. The controller18 may further include a dosimetry calculator 28 to calculate the dosageneeded for a particular patient based on the application type and otherapplication site information input into the controller by a physician.In one form, the controller 18 further includes an imaging system forimaging the application site. The imaging system gathers applicationsite information based on the images of the application site andtransfers the gathered information to the dosimetry calculator 28 fordosage calculation. A physician also can manually calculate and inputinformation gathered from the images to the controller 18.

As shown in FIG. 2, the controller may further include a control panel30 through which, a physician can control the therapeutic systemmanually. The therapeutic system 10 also can be controlled by acomputer, which has a control platform, for example, a WINDOWS™ basedplatform. The parameters such as pulse intensity, pulse width, pulserepetition rate of the optical radiation can be controlled through boththe computer and the control panel 30.

FIGS. 3 a-3 d show different patterns of the optical radiation that canbe delivered from the therapeutic system to the application site. Theoptical radiation can be delivered in one wavelength range only, forexample, in the first wavelength range of 850 nm to 900 nm, or in therange of 865 nm to 875 nm, or in the second wavelength range of 905 nmto 945 nm, or in the range of 925 nm to 935 nm, as shown in FIG. 3 a.The radiation in the first wavelength range and the radiation in thesecond wavelength range also can be multiplexed by a multiplex systeminstalled in the optical radiation generation device 12 and delivered tothe application site in a multiplexed form, as shown in FIG. 3 b. In analternative form, the radiation in the first wavelength range and theradiation in the second wavelength range can be applied to theapplication site simultaneously without passing through a multiplexsystem. FIG. 3 c shows that the optical radiation can be delivered in anintermission-alternating manner, for example, a first pulse in the firstwavelength range, a second pulse in the second wavelength range, a thirdpulse in the first wavelength range again, and a fourth pulse in thesecond wavelength range again, and so on. The interval can be CW(Continuous Wave), one pulse as shown in FIG. 3 c, or two or more pulses(not shown). FIG. 3 d shows another pattern in which the applicationsite is first treated by radiation in one of the two wavelength ranges,for example, the first wavelength range, and then treated by radiationin the other wavelength range. The treatment pattern can be determinedby the physician based on the type, and other information of theapplication site.

The following examples are intended to further illustrate certainexemplary embodiments of the disclosure, and are not intended to limitthe scope of the disclosure.

EXAMPLE I NIMELS Dosimetry Calculations

As discussed in more details supra NIMELS parameters include the averagesingle or additive output power of the laser diodes, and the wavelengths(870 nm and 930 nm) of the diodes. This information, combined with thearea of the laser beam or beams (cm²) at the target site, provide theinitial set of information which may be used to calculate effective andsafe irradiation protocols according to the disclosure.

The power density of a given laser measures the potential effect ofNIMELS at the target site. Power density is a function of any givenlaser output power and beam area, and may be calculated with thefollowing equations:

For a single wavelength:

${\text{1)}\mspace{14mu} {Power}\mspace{14mu} {Density}\mspace{14mu} \left( {W\text{/}{cm}^{2}} \right)} = \frac{{Laser}\mspace{14mu} {Output}\mspace{14mu} {Power}}{{Beam}\mspace{14mu} {Diameter}\mspace{14mu} \left( {cm}^{2} \right)}$

For dual wavelength treatments:

${\text{2)}\mspace{14mu} {Power}\mspace{14mu} {Density}\mspace{14mu} \left( {W\text{/}{cm}^{2}} \right)} = {\frac{{Laser}\mspace{14mu} (1)\mspace{14mu} {Output}\mspace{14mu} {Power}}{{Beam}\mspace{14mu} {Diameter}\mspace{14mu} \left( {cm}^{2} \right)} + \frac{{Laser}\mspace{14mu} (2)\mspace{14mu} {Output}\mspace{14mu} {Power}}{{Beam}\mspace{14mu} {Diameter}\mspace{14mu} \left( {cm}^{2} \right)}}$

Beam area can be calculated by either:

Beam Area (cm²)=Diameter (cm)²*0.7854 or Beam Area (cm²)=Pi* Radius(cm)²   3)

The total photonic energy delivered into the tissue by one NIMELS laserdiode system operating at a particular output power over a certainperiod is measured in Joules, and is calculated as follows:

Total Energy (Joules)=Laser Output Power (Watts)*Time (Secs.)   4)

The total photonic energy delivered into the tissue by both NIMELS laserdiodes systems (both wavelengths) at the same time, at particular outputpowers over a certain period, is measured in Joules, and is calculatedas follows:

Total Energy (Joules)=[Laser(1) Output Power (Watts)*Time (Secs)]+[Laser(2) Output Power (Watts)*Time(Secs)]  5)

In practice, it is useful (but not necessary) to know the distributionand allocation of the total energy over the irradiation treatment area,in order to correctly measure dosage for maximal NIMELS beneficialresponse. Total energy distribution may be measured as energy density(Joules/cm²). As discussed infra, for a given wavelength of light,energy density is the most important factor in determining the tissuereaction. Energy density for one NIMELS wavelength may be derived asfollows:

${\text{6)}\mspace{14mu} {Energy}\mspace{14mu} {Density}\mspace{14mu} \left( {{Joules}\text{/}{cm}^{2}} \right)} = \frac{{Laser}\mspace{14mu} {Output}\mspace{14mu} {power}\mspace{14mu} ({Watts})*{Time}\mspace{14mu} ({secs})}{{Beam}\mspace{14mu} {Area}\mspace{14mu} \left( {cm}^{2} \right)}$7)  Energy  Density  (Joules/cm²) = Power  Density  (W/cm²) * Time  (secs)

When two NIMELS wavelengths are being used, the energy density may bederived as follows:

${{\text{8)}\mspace{14mu} {Energy}\mspace{14mu} {Density}\mspace{14mu} \left( {{Joules}\text{/}{cm}^{2}} \right)} = {\frac{{Laser}\mspace{14mu} (1)\mspace{14mu} {Output}\mspace{14mu} {power}\mspace{14mu} ({Watts})*{Time}\mspace{14mu} ({secs})}{{Beam}\mspace{14mu} {Area}\mspace{14mu} \left( {cm}^{2} \right)} + {\frac{{Laser}\mspace{14mu} (2)\mspace{14mu} {Output}\mspace{14mu} {power}\mspace{14mu} ({Watts})*{Time}\mspace{14mu} ({secs})}{{Beam}\mspace{14mu} {Area}\mspace{14mu} \left( {cm}^{2} \right)}\mspace{14mu} {or}}}},{{\text{9)}\mspace{14mu} {Energy}\mspace{14mu} {Density}\mspace{14mu} \left( {{Joule}\text{/}{cm}\; 2} \right)} = {{{Power}\mspace{14mu} {Density}\mspace{14mu} (1)\mspace{14mu} \left( {W\text{/}{cm}^{2}} \right)*{Time}\mspace{14mu} ({Secs})} + {{Power}\mspace{14mu} {Density}\mspace{14mu} (2)\mspace{14mu} \left( {W\text{/}{cm}^{2}} \right)*{Time}\mspace{14mu} ({Secs})}}}$

To calculate the treatment time for a particular dosage, a user may useeither the energy density (J/cm²) or energy (J), as well as the outputpower (W), and beam area (cm²) using either one of the followingequations:

${\text{10)}\mspace{14mu} {Treatment}\mspace{14mu} {Time}\mspace{14mu} ({seconds})} = \frac{{Energy}\mspace{14mu} {Density}\mspace{14mu} \left( {{Joules}\text{/}{cm}^{2}} \right)}{{Output}\mspace{14mu} {power}\mspace{14mu} {Density}\mspace{14mu} \left( {W\text{/}{cm}^{2}} \right)}$${\text{11)}\mspace{14mu} {Treatment}\mspace{14mu} {Time}\mspace{14mu} ({seconds})} = \frac{{Energy}\mspace{14mu} ({Joules})}{{Laser}\mspace{14mu} {Output}\mspace{14mu} {Power}\mspace{14mu} ({Watts})}$

Because dosimetry calculations such as those exemplified in this Examplecan become burdensome, the therapeutic system may also include acomputer database storing all researched treatment possibilities anddosimetries. The computer (a dosimetry and parameter calculator) in thecontroller is preprogrammed with algorithms based on the above-describedformulas, so that any operator can easily retrieve the data andparameters on the screen, and input additional necessary data (such as:spot size, total energy desired, time and pulse width of eachwavelength, tissue being irradiated, bacteria being irradiated) alongwith any other necessary information, so that any and all algorithms andcalculations necessary for favorable treatment outcomes can be generatedby the dosimetry and parameter calculator and hence run the laser.

The following examples describe selected experiments showing the abilityof the NIMELS approach to impact upon the viability of various commonlyfound microorganisms at the wavelengths as described herein. Themicroorganisms exemplified include E. coli K-12, multi-drug resistant E.coli, Staphylococcus aureus, Methicillin-resistant S. aureus, Candidaalbicans, and Trichophyton rubrum.

In summary, when the bacterial cultures were exposed to the NIMELSlaser, the bacterial kill rate (as measured by counting Colony FormingUnits or CFU on post-treatment culture plates) ranged from 93.7%(multi-drug resistant E. coli) to 100% (all other bacteria and fungi).

EXAMPLE II Bacterial Methods: NIMELS Treatment Parameters for In VitroE. coli Targeting

The following parameters illustrate the methods according to thedisclosure as applied to E. coli, at final temperatures well below thoseassociated in the literature with thermal damage.

A. Experiment Materials and Methods for E. coli K-12:

E. coli K1.2 liquid cultures were grown in Luria Bertani (LB) medium (25g/L). Plates contained 35 mL of LB plate medium (25 g/L LB, 15 g/Lbacteriological agar). Cultures dilutions were performed usingphosphate-buffered saline (PBS). All protocols and manipulations wereperformed using sterile techniques.

B. Growth Kinetics

Drawing from a seed culture, multiple 50 mL LB cultures were inoculatedand grown at 37° C. overnight. The next morning, the healthiest culturewas chosen and used to inoculate 5% into 50 mL LB at 37° C. and theO.D.₆₀₀ was monitored over time taking measurements every 30 to 45minutes until the culture was in stationary phase.

C. Master Stock Production

Starting with a culture in log phase (O.D.₆₀₀ approximately 0.75), 10 mLwere placed at 4° C. 10 mL of 50% glycerol were added and then this wasaliquoted to 20 cryovials and snap frozen in liquid nitrogen. Thecryovials were then stored at −80° C.

D. Liquid Cultures

Liquid cultures of E. coli K12 were set up as described previously. Analiquot of 100 μL was removed from the subculture and serially dilutedto 1:1200 in PBS. This dilution was allowed to incubate at roomtemperature approximately 2 hours or until no further increase inO.D.₆₀₀ was observed in order to ensure that the cells in the PBSsuspension would reach a static state (growth) with no significantdoubling and a relatively consistent number of cells could be aliquotedfurther for testing.

Once it was determined that the K12 dilution was in a static state, 2 mLof this suspension were aliquoted into selected wells of 24-well tissueculture plates for selected NIMELS experiments at given dosimetryparameters. The plates were incubated at room temperature until readyfor use (approximately 2 hrs).

Following laser treatments, 100 μl was removed from each well andserially diluted to 1:1000 resulting in a final dilution of 1:12×10⁵ ofinitial K12 culture. Aliquots of 3×200 L of each final dilution werespread onto separate plates in triplicate. The plates were thenincubated at 37° C. for approximately 16 hours. Manual colony countswere performed and recorded. A digital photograph of each plate was alsotaken.

Similar cell culture and kinetic protocols were performed for all NIMELSirradiation tests with S. aureus and C. albicans in vitro tests. Hence,for example, C. albicans ATCC 14053 liquid cultures were grown in YMmedium (21 g/L, Difco) medium at 37° C. A standardized suspension wasaliquoted into selected wells in a 24-well tissue culture plate.Following laser treatments, 100 μL was removed from each well andserially diluted to 1:1000 resulting in a final dilution of 1:5×10⁵ ofinitial culture. 3×100 μL of each final dilution were spread ontoseparate plates. The plates were then incubated at 37° C. forapproximately 16-20 hours. Manual colony counts were performed andrecorded. A digital photograph of each plate was also taken.

T. rubrum ATCC 52022 liquid cultures were grown in peptone-dextrose (PD)medium at 37° C. A standardized suspension was aliquoted into selectedwells in a 24-well tissue culture plate. Following laser treatments,3×100 μL aliquots were removed from each well and spread onto separateplates. The plates were then incubated at 37° C. for approximately 91hours. Manual colony counts were performed and recorded after 66 hoursand 91 hours of incubation. While control wells all grew the organism,100% of laser-treated wells as described herein had no growth. A digitalphotograph of each plate was also taken.

Thermal tests performed on PBS solution, starting from room temperature.10 Watts of NIMELS laser energy is available for use in a 12 minutelasing cycle, before the temperature of the system is raised close tothe critical threshold of 44° C.

TABLE II Time Temperature measurements for In Vitro NIMELS DosimetriesBEAM SPOT ENERGY 1.5 CM DENSITY POWER NIMEL DIAMETER TOTAL (RADIANTDENSITY OUTPUT OVERLAP AREA TREATMENT ENERGY EXPOSURE) (IRRADIANCE)TEMPERATURE TEMP POWER (W) (CM²) TIME (SEC) (JOULES) (J/CM²) (W/CM²)START FINISH Plate 1-N - 1.76 720 4320 2448 3.40 20.5° C. 34.0° C. 3.0 +3.0 = 6.0 W Plate 2-N - 1.76 720 5040 2858 3.97 20.7° C. 36.5° C. 3.5 +3.5 = 7.0 W Plate 3-N - 4.0 + 1.76 720 5760 3268 4.54 21.0° C. 38.5° C.4.0 = 8.0 W Plate 4-N - 4.5 + 1.76 720 6480 3679 5.11  2.0° C. 41.0° C.4.5 = 9.0 W Plate 5-N - 5.0 + 1.76 720 7200 4089 5.68 21.0° C. 40.5° C.5.0 = 10. W Plate 6-N - 5.5 + 1.76 720 7920 4500 6.25 21.0° C. 46.0° C.5.5 = 11 W Plate 7-N - 7.0 + 1.76 360 5040 2863 7.95 21.0° C. 47.0° C.7.0 = 14.0 W Plate 8-N - 7.5 + 1.76 360 5400 3068 8.52 21.7° C. 47.2° C.7.5 = 15 W

EXAMPLE III Dosimetry Values for NIMELS Laser Wavelength 930 nm for E.coli In Vitro Targeting

The instant experiment shows that the NIMELS single wavelength λ=930 nmwas associated with quantitatable antibacterial efficacy against E. coliin vitro within safe thermal parameters for mammalian tissues.

Experimental data in vitro demonstrates that if the threshold of totalenergy into the system with 930 nm alone of 5400 J and an energy densityof 3056 J/cm² is met in 25% less time, 100% antibacterial efficacy isstill achieved.

TABLE III Sub-thermal NIMELS (λ = 930) Dosimetry for In Vitro E. coliTargeting OUTPUT TOTAL ENERGY POWER POWER BEAM SPOT TIME ENERGY DENSITYDENSITY E-COLI KILL (W) (CM) (SEC.) JOULES (J/CM²) (W/CM²) PERCENTAGE7.0 1.5 720 5040 2852 3.96 40.2% 8.0 1.5 720 5760 3259 4.53 100.0% 10.01.5 540 5400 3056 5.66 100.0%

Experimental data in vitro also demonstrated that treatments using asingle energy with λ=930 nm had antibacterial efficacy against thebacterial species S. aureus in vitro within safe thermal parameters formammalian tissues.

It is also believed that if the threshold of total energy into thesystem of 5400 J and an Energy Density of 3056 J/cm² is met in 25% lesstime with S. aureus and other bacterial species, that 100% antibacterialefficacy will still be achieved.

TABLE IV Sub-thermal NIMELS (λ = 930) Dosimetry for In Vitro S. aureusTargeting OUTPUT TOTAL ENERGY POWER POWER BEAM ENERGY DENSITY DENSITY SAUREUS KILL (W) SPOT (CM) TIME (SEC) JOULES (J/CM²) (W/CM²) PERCENTAGE7.0 1.5 720 5040 2852 3.96 24.1% 8.0 1.5 720 5760 3259 4.53 100.0%

Experimental data in vitro also showed that the NIMELS single wavelengthof λ=930 nm had anti-fungal efficacy against C. albicans in vitro atranges within safe thermal parameters for mammalian tissues.

It is also believed that if the threshold of total energy into thesystem of 5400 J and an energy density of 3056 J/cm² is met in 25% lesstime, that 100% antifungal efficacy will still be achieved, See alsoFIG. 3.

TABLE V Sub-thermal NIMELS (λ = 930) Dosimetry for In Vitro C. albicansTargeting CANDIDA OUTPUT TOTAL ENERGY POWER ALBICANS POWER BEAM TIMEENERGY DENSITY DENSITY KILL (W) SPOT (CM) (SEC.) JOULES (J/CM²) (W/CM²)PERCENTAGE 8.0 1.5 720 5760 3259 4.53 100.0% 9.0 1.5 720 6840 3681 5.11100.0%

EXAMPLE IV Dosimetry Values for NIMELS Laser Wavelength 870 nm In Vitro

Experimental data in vitro also demonstrated that no significant killwas achieved up to a total energy of 7200 J, and energy density of 4074J/cm² and a power density of 5.66 0 W/cm² with the wavelength of 870 nmalone against E. coli.

TABLE VI E. coli Studies- Single wavelength λ = 870 nm OUTPUT BEAM TOTALENERGY POWER DIFFERENCE POWER SPOT TIME ENERGY DENSITY DENSITY CONTROLNIMELS CONTROL- E. COLI KILL (W) (CM) (SEC.) JOULES (J/CM²) (W/CM²) CFUsCFUs NIMEL PERCENTAGE 6.0 1.5 720 4320 2445 3.40 90 95 (5) −5.6% 7.0 1.5720 5040 2852 3.96 94 94 0 0.0% 8.0 1.5 720 5760 3259 4.53 93 118 (25) −26.9% 9.0 1.5 720 6480 3667 5.09 113 112 1 0.9% 10.0 1.5 720 7200 40745.66 103 111 (8) −7.8% 10.0 1.5 540 5400 3056 5.66 120 101 19  15.8%Comparable results using radiation having λ=870 nm alone were alsoobserved with S. aureus.

EXAMPLE V NIMELS Unique Alternating Synergistic Effect Between 870 nmand 930 nm Optical Energies

Experimental data in vitro also demonstrated that there is an additiveeffect between the two NIMELS wavelengths (λ=870 nm and 930 nm) whenthey are alternated (870 nm before 930 nm). The presence of the 870 nmNIMELS wavelength as a first irradiance has been found to enhance theeffect of the antibacterial efficacy of the second 930 nm NIMELSwavelength irradiance.

Experimental data in vitro demonstrates that this synergistic effect(combining the 870 nm wavelength to the 930 nm wavelength) allows forthe 930 nm optical energy to be reduced. As shown hereinafter, theoptical energy was reduced to approximately ⅓ of the total energy andenergy density required for NIMELS 100% E. coli antibacterial efficacy,when the (870 nm before 930 nm) wavelengths are combined in analternating manner.

Experimental data in vitro also demonstrates that this synergisticmechanism can allow for the 930 nm optical energy (total energy andenergy density) to be reduced to approximately ½ of the total energydensity necessary for NIMELS 100% E. coli antibacterial efficacy ifequal amounts of 870 nm optical energy are added to the system beforethe 930 nm energy at 20% higher power densities.

TABLE VII E. coli data from Alternating NIMELS Wavelengths OUTPUT POWERPOWER SPOT TOTAL ENERGY ENERGY DENSITY DENSITY E. COLI KILL (W) (CM)TIME (SEC.) JOULES (J/CM²) (W/CM²) PERCENTAGE 8 W/8 W 1.5 540/1804320/1440 = 2445/815 = 4.53/4.53 100.0% 12 min. 5760 3529 10 W/10 W 1.5240/240 2400/2400 = 1358/1358 = 5.66/5.66 100.0%  8 min. 4800 2716

This synergistic ability is significant to human tissue safety, as the930 nm optical energy, heats up a system at a greater rate than the 870nm optical energy, and it is beneficial to a mammalian system to producethe least amount of heat possible during treatment.

It is also believed that if the NIMELS optical energies (870 nm and 930nm) are alternated in the above manner with other bacterial species,that the 100% antibacterial effect will be essentially the same.

Experimental data in vitro also demonstrates that there is also anadditive effect between the two NIMELS wavelengths (870 nm and 930 nm)when they are alternated (870 nm before 930 nm) while irradiating fungi.The presence of the 870 nm NIMELS wavelength as a first irradiancemathematically enhances the effect of the anti-fungal efficacy of thesecond 930 nm NIMELS wavelength irradiance.

Experimental data in vitro (see, table infra) demonstrates that thissynergistic mechanism can allow for the 930 nm optical energy (totalenergy and energy density) to be reduced to approximately ½ of the totalenergy density necessary for NIMELS 100% antifungal efficacy if equalamounts of 870 rim optical energy is added to the system before the 930nm energy at 20% higher power densities than is required for bacterialspecies antibacterial efficacy.

TABLE VIII C. albicans Data from Alternating NIMEL Wavelengths CANDIDAOUTPUT POWER ALBICANS POWER SPOT TOTAL ENERGY ENERGY DENSITY DENSITYKILL (W) (CM) TIME (SEC) JOULES (J/CM²) (W/CM²) PERCENTAGE 10 W/10 W 1.5240/240 2400/2400 = 1358/1358 = 5.66/5.66 100.0%* 8 min 4800 2716

This synergistic effect is significant to human tissue safety, as the930 nm optical energy, heats up a system at a greater rate than the 870nm optical energy, and it is beneficial to a mammalian system to producethe least amount of heat possible during treatment.

It is also believed that if the NIMELS optical energies (870 nm and 930nm) are alternated in the above manner with other fungi species, thatthe 100% anti-fungal effect will be essentially the same.

EXAMPLE VI NIMELS Unique Simultaneous Synergistic Effect Between λ=870nm and λ=930 nm Optical Energies

Experimental data in vitro also demonstrates that there is an additiveeffect between the two NIMELS wavelengths (870 nm and 930 nm) when theyare used simultaneously (870 nm combined with 930 nm). The presence ofthe 870 nm NIMELS wavelength and the 930 nm NIMELS wavelength as asimultaneous irradiance absolutely enhances the effect of theantibacterial efficacy of the NIMELS system.

In vitro experimental data (see, for example, Tables IX and X below)demonstrated that by combining λ=870 nm and λ=930 nm (in this example,used simultaneously) effectively reduces the 930 nm optical energy anddensity by about half of the total energy and energy density requiredwhen using a single treatment according to the disclosure.

TABLE IX E. coli data from Combined NIMEL Wavelengths OUTPUT POWER (W)BEAM TOTAL 870 NM/ SPOT ENERGY ENERGY DENSITY POWER DENSITY E-COLI KILL930 NM (CM) TIME (SEC) JOULES (J/CM²) (W/CM²) PERCENTAGE 5 W + 5 W = 101.5 720 3600 (×2) = 2037 (×2) = 5.66 100% 7200 4074

TABLE X S. aureus data from Combined NIMELS Wavelengths OUTPUT POWER (W)BEAM TOTAL 870 NM/ SPOT ENERGY ENERGY DENSITY POWER DENSITY S. AUREUSKILL 930 NM (CM) TIME (SEC) JOULES (J/CM²) (W/CM²) PERCENTAGE 5 W + 5 W= 10 W 1.5 720 3600 (×2) = 2037 (×2) = 5.66 98.5% 7200 4074 55 W + 5.5 =11 W 1.5 720 3960 (×2) = 2241 (×2) = 6.22  100% 7920 4482

This simultaneous synergistic ability is significant to human tissuesafety, as the 930 nm optical energy, heats up a system at a greaterrate than the 870 nm optical energy, and it is beneficial to a mammaliansystem to produce the least amount of heat possible during treatment.

It is now understood that if the NIMELS optical energies (870 nm and 930nm) are used simultaneously in the above manner with other bacterialspecies, that the 100% antibacterial effect will be essentially thesame. See FIGS. 4 and 5.

Experimental data in vitro also demonstrates that there is an additiveeffect between the two NIMELS wavelengths (870 nm and 930 nm) when theyare used simultaneously on fungi. The presence of the 870 nm NIMELSwavelength and the 930 nm NIMELS wavelength as a simultaneous irradiancehave been found to enhance the effect of the anti-fungal efficacy of theNIMELS system.

Experimental data in vitro (see Table X) demonstrates that thissynergistic effect (connecting the 870 nm wavelength to the 930 nmwavelength for simultaneous irradiation) allows for the 930 nm opticalenergy to be reduced to approximately ½ of the total energy and energydensity required for NIMELS 100% C. albicans anti-fungal efficacy, whenthe (870 nm before 930 nm) wavelengths are combined in a simultaneousmanner.

TABLE XI Candida albicans from Combined NIMELS Wavelengths OUTPUT POWER(W) BEAM TOTAL C. ALBICANS 870 NM/ SPOT ENERGY ENERGY DENSITY POWERDENSITY KILL 930 NM (CM) TIME (SEC) JOULES (J/CM²) (W/CM²) PERCENTAGE 5W + 5 W = 10 1.5 720 3600 (×2) = 2037 (×2) = 4074 5.66 100% 7200

Thus NIMELS wavelengths (λ=870 nm and 930 nm) may be used to achieveantibacterial and anti-fungal efficacy in an alternating mode orsimultaneously or in any combination of such modes thereby reducing theexposure at the λ=930 associated with temperature increases which arepreferably minimized.

Experimental data in vitro also demonstrates that when E. coli isirradiated alone with a (control) wavelength of λ=830 nm, at thefollowing parameters (see Table XI), the control 830 nm laser producedzero antibacterial efficacy for 12 minutes irradiation cycles, atidentical parameters to the minimum NIMELS dosimetry associated with100% antibacterial and anti-fungal efficacy with radiation of λ=930 nm.

TABLE XII E. coli Single Wavelength λ = 830 nm OUTPUT BEAM TOTAL ENERGYPOWER POWER SPOT TIME ENERGY DENSITY DENSITY (W) (CM) (SEC.) JOULES(J/CM²) (W/CM²) 8.0 1.5 720 5760 3259 4.53 9.0 1.5 720 6480 3667 5.09

Experimental data in vitro also demonstrates that when applied at safethermal dosimetries, there is little additive effect when using radianceof λ=830 nm in combination with λ=930 nm. The presence of the 830 nmcontrol wavelength as a first irradiance, is far inferior to theenhancement effect of the 870 nm NIMELS wavelength in producingsynergistic antibacterial efficacy with the second 930 nm NIMELSwavelength.

TABLE XIII E. coli data from Substituted alternating 830 nm controlWavelength OUTPUT POWER (W) 830 NM/ BEAM SPOT TIME TOTAL ENERGY ENERGYDENSITY POWER DENSITY E. COLI KILL 930 NM (CM) (SEC) JOULES (J/CM²)(W/CM²) PERCENTAGE  8 W/8 W 1.5 540/180 4320/1440 = 5760 2445/815 = 35294.53/4.53 0% 12 min 10 W/10 W 1.5 240/240 2400/2400 = 4800 1358/1358 =2716 5.66/5.66 65%  8 min

Experimental data in vitro also demonstrates that when applied at safethermal dosimetries, there is less additive effect with the 830 nmwavelength, and the NIMELS 930 nm wavelength when they are usedsimultaneously. In fact, experimental data in vitro demonstrates that17% less total energy, 17% less energy density, and 17% less powerdensity is required to achieve 100% E. coli antibacterial efficacy when870 nm is combined simultaneously with 930 nm, vs. the commerciallyavailable 830 nm. This again substantially reduces heat and harm to thein vivo system being treated with the NIMELS wavelengths.

TABLE XIV E. coli data from Substituted Simultaneous 830 nm controlWavelength OUTPUT POWER (W) BEAM TOTAL 830 NM/ SPOT TIME ENERGY ENERGYDENSITY POWER DENSITY E-COLI KILL 930 NM (CM) (SEC) JOULES (J/CM²)(W/CM²) PERCENTAGE 5 W + 5 W = 10 1.5 720 3600 (×2) = 2037 (×2) = 40745.66 91% 7200 5.5 W + 5.5 = 1.5 720 3960 (×2) = 2250 (×2) = 4500 6.2590% 11 W 7920 6 W + 6 W = 1.5 720 3960 (×2) f = 2454 (×2) = 6.81* 100%12 W 8640* 4909*

Amount of Bacteria Killed:

In vitro data also showed that the NIMELS laser system in vitro iseffective (within thermal tolerances) against solutions of bacteriacontaining 2,000,000 (2×10⁶) Colony Forming Units (CFU's) of E. coli andS. aureus. This is a 2× increase over what is typically seen in a 1 gmsample of infected human ulcer tissue. Brown et al, reported thatmicrobial cells in 75% of the diabetic patients tested were all at least100,000 CFU/gm, and in 37.5% of the patients, quantities of microbialcells were greater than 1,000,000 (1×10⁶)CFU (see, Brown et al., OstomyWound Management, 401:47, issue 10, (2001) the entire teachings of whichare incorporated herein by reference in their entirety).

Thermal Parameters:

Experimental data in vitro also demonstrates that the NIMELS lasersystem can accomplish 100% antibacterial and anti-fungal efficacy withinsafe thermal tolerances for human tissues. See FIG. 6.

EXAMPLE VII The Effects of Lower Temperatures on NIMELS

Dewhirst et al., Internat. J. of Hyperthermia, 19(3):267-294 reportedthe effects of lower temperature on bacteria; the entire teachings ofwhich are incorporated herein by reference in their entirety.

Cooling of Bacterial Species:

Experimental data in vitro also demonstrated that by substantiallylowering the starting temperature of bacterial samples to 4° C. for twohours in PBS before lasing cycle, that optical antibacterial efficacywas not achieved at any currently reproducible antibacterial energieswith the NIMELS laser system.

The most probable explanation is that the bacterial cells were in“metabolic stasis” and that little or no radical oxygen was producedwithout active metabolism occurring in the cells. This data indicatesthat NIMELS may be affecting respiratory centers and cell membranes ofthe targeted microorganisms.

The postulated mechanism is that the 870 nm energy effects thecytochromes by speeding up oxidative phosphorylation while the 930 nmenergy disrupts cell membranes and hence produces singlet oxygen viauncoupling the electron transport system, and not allowing the terminalO₂ molecule to be reduced.

EXAMPLE VIII Trychophyton Rubrum

TABLE XV NIMELS T. rubrum Tests Alternating Wavelengths OUTPUT POWER (W)870 NM/ BEAM TOTAL ENERGY ENERGY DENSITY POWER DENSITY EXP. No. 930 NMSPOT (CM) TIME (SEC.) JOULES (J/CM²) (W/CM²) 1 8 W/8 W 1.5 540/1804320/1440 = 2445/815 = 4.53/4.53 12 min. 5760 3529 2 10 W/10 W 1.5240/240 2400/2400 = 1358/1358 = 5.66/5.66  8 min. 4800 2716 ExperimentNo. 1 = Minimal Effect Experiment No. 2 = 100% Kill in all plates

TABLE XVI NIMELS T. rubrum -- Simultaneous Wavelengths OUTPUT POWER (W)EX Λ = 870 NM & BEAM TOTAL ENERGY ENERGY DENSITY POWER DENSITY NO. Λ =930 NM SPOT (CM) TIME (SEC.) JOULES (J/CM²) (W/CM²) 3 5 + 5 = 10 1.5 7203600 (×2) = 7200 2037 (×2) = 5.66 12 min. 4074 4 5.5 W + 5.5 W = 1.5 7203960 (×2) = 7920 2250 (×2) = 4500 6.25 11 W 5 6 W + 6 W = 1.5 720 3960(×2) = 8640 2454 (×2) = 4909 6.81 12 W Experiments Nos. 3, 4, and 5 =100% Kill in all plates

TABLE XVII NIMELS T. rubrum - Single Wavelength BEAM TOTAL EXP NO OUTPUTSPOT ENERGY ENERGY DENSITY POWER DENSITY λ = 930 POWER (W) (CM) TIME(SEC.) JOULES (J/CM²) (W/CM²) 6 8.0 1.5 720 5760 3259 4.53 7 9.0 1.5 7206840 3681 5.11 Experiments Nos. 6 and 7 = 100% Kill in all plates

TABLE XVIII Control T. rubrum -- 830 nm/930 nm Alternating EXPERIMENTNO. OUTPUT λ 830 & POWER BEAM TOTAL ENERGY ENERGY DENSITY POWER DENSITYλ = 930 (W) SPOT (CM) TIME (MIN.) JOULES (J/CM²) (W/CM²) 8 8 W/8 W 1.5540/180 4320/1440 = 5760 2445/815 = 3529 4.53/4.53 12 min 9 10 W/10 W1.5 240/240 2400/2400 = 4800 1358/1358 = 2716 5.66/5.66  8 minExperiment No. 8 = No Effect Experiment No. 9 = 100% Kill

TABLE XIX In Vitro Targeting of T. rubrum using λ = 830 nm and 930 nmBEAM TOTAL ENERGY POWER OUTPUT SPOT TIME ENERGY DENSITY DENSITY POWER(W) (CM) (SEC.) JOULES (J/CM²) (W/CM²) 5 + 5 = 10 1.5 720 3600 (×2) =2037 (×2) = 5.66 7200 4074

Treatments as described in the above Table XVIII resulted in 100% kill.

EXAMPLE IX Onychomycosis Treatment Evaluation

This example is provided to illustrate how a practitioner's evaluationaids and informs in evaluating whether to increase, reduce or continue aparticular treatment dose, mode of irradiation. Making reference to FIG.8, the healthy nail plate is hard and translucent, and is composed ofdead keratin. The plate is surrounded by the perionychium, whichconsists of proximal and lateral nail folds, and the hyponychium, thearea beneath the free edge of the nail. The nail bed is beneath the nailplate and contains the blood vessels and nerves. Contained in the nailbed is the germinal matrix, which produces most of the nails keratinizedvolume, and the sterile matrix. This matrix is the “root” of the nail,and its most distal portion is visible on many nails as the half-moonshaped structure called the lunula.

FIG. 9 shows the diagram of a typical onychomycosis patient's nailevidencing the effectiveness of the treatment by the presence of healthynail growth. The practitioner will recognize that the clean and“unifected” portion of the newly growing nail plate (proximal to thegerminal matrix, eponychium and lunula) will not automatically need tobe irradiated in subsequent treatments. Hence, the irradiation spotshould potentially be aimed preferentially or only at the diseasedareas, that are still impregnated with the pathogen(s).

In certain instances nails infected with onychomycosis are inherently“thicker” (because of dystrophic growth) or “colored” (because of thechroma produced by the fungal pathogen) (see, FIG. 10) and may require alonger lasing time (higher energy density) to penetrate through the nailplate to the infected areas of the bed (sterile matrix and germinalmatrix) and nail fold lunula growing out under the Eponychium). As shownin FIG. 10, paronychial infections can develop when a disruption occursbetween the seal of the proximal nail fold and the nail plate thatallows a portal of entry for invading organisms. Chronic paronychia as arule, causes swollen, red, tender and boggy nail folds where thesymptoms of the disease present for six weeks or longer and areconcomitant with long term onychomycosis.

FIG. 15 is a composite showing the improvement over time in theappearance of the nail of a typical onychomycosis patient treatedaccording to the methods of the disclosure.

As shown in FIG. 11, in patients with concurrent chronic paronychia, the“spot size” of the laser treatment area should be expanded to cover theinfected paronychial regions to be sure that all of the pathogeninfected areas of the nail complex are treated with the NIMELS laser.

In certain cases, onychomycosis patients may have different discreteareas of the nail infected with a pathogen, and other areas that arecompletely clean where the healthy portion of the nail plate is stillhard and translucent (ref. to FIG. 11). This may be in a vertical orhorizontal pattern and can reach to and beyond the lunula growing outunder the eponychium. In these cases, the practitioner will recognizethat the clean and “unifected” portion of the nail plate will notautomatically need to be irradiated, and the spot size and concominentlaser dosimetry will be adjusted accordingly to allow successfultreatment without damaging any part of the healthy nail complex. Also,the healthy part of the nail could be covered with an opaque substanceto allow for a larger irradiation spot from the laser, if the geometryof the infected part of the nail could not be adequately treated withsimply a “smaller spot”.

EXAMPLE X Reciprocal Progression Analysis for In Vivo NIMELS Therapywith Output Power of Laser fixed at 3.0 Watts Combined: both 870 and 930at 1.5 W

To illustrate typical analysis performed for in vivo therapy, thefollowing example assumed the use of a laser with a power output of 3 Wto emit energy with λ=870 and 930 nm.

TABLE XX Dual Wavelengths λ = 870 and 930 nm. BEAM TOTAL ENERGY POWEROUTPUT SPOT AREA OF TIME ENERGY DENSITY DENSITY POWER (W) (CM) SPOT(CM2) (SEC) JOULES (J/CM²) (W/CM²) 3.0 1.2 1.13 154 462 408 2.65 3.0 1.31.33 180 540 407 2.26 3.0 1.4 1.54 210 630 409 1.95 3.0 1.5 1.77 240 720407 1.70 3.0 1.6 2.01 272 816 406 1.49 3.0 1.7 2.27 309 927 408 1.32 3.01.8 2.54 345 1035 407 1.18 3.0 1.9 2.84 382 1146 404 1.06 3.0 2 3.14 4281284 409 0.95 3.0 2.1 3.46 472 1416 409 0.87 3.0 2.2 3.80 514 1542 4060.79

In this context, Tn=409 (Energy density)/Power Density. FIG. 14, showsderived values for a given spot-size (1.2-2.2 cm diameter). Treatmenttime for NIMELS therapy was derived dividing an Energy Density of 409J/cm² by the Power Density, at a laser output power of 3.0 Watts. Hence,NIMELS (Time) Factor=Tn=409/Power Density.

EXAMPLE XI Reciprocal Progression Analysis for In Vivo NIMELS Therapywith Output Power of Laser fixed at 3.0 Watts and Wavelength at 930 nm

To illustrate typical analysis performed for in vivo therapy, thefollowing example assumed the use of a laser with a power output of 3 Wto emit energy with λ=930 nm.

TABLE XXI Single Wavelength λ = 930 nm. BEAM TOTAL ENERGY POWER OUTPUTSPOT AREA OF ENERGY DENSITY DENSITY POWER (W) (CM) SPOT (CM²) TIME (SEC)JOULES (J/CM²) (W/CM²) 3.0 1.2 1.13 77 231 204 2.65 3.0 1.3 1.33 90 270203 2.26 3.0 1.4 1.54 105 315 205 1.95 3.0 1.5 1.77 120 360 204 1.70 3.01.6 2.01 137 411 204 1.49 3.0 1.7 2.27 155 465 205 1.32 3.0 1.8 2.54 172516 203 1.18 3.0 1.9 2.84 194 582 205 1.06 3.0 2 3.14 214 642 204 0.953.0 2.1 3.46 233 699 202 0.87 3.0 2.2 3.80 256 768 202 0.79

On the basis of the observed value (see, data above in Table XXI) it isfound that Tn=205 (energy density)/power density. Hence, within thegiven spot-size parameters (1.2-2.2 cm diameter), treatment time forNIMELS therapy can be simply derived dividing an energy density of 205J/cm² by the power density, at a laser output power of 3.0 Watts (seeFIG. 13). Hence, NIMELS (Time) Factor=Tn=205/Power Density.

This novel algorithm for NIMELS dosimetry calculations concerns thequantification of a known and constant NIMELS threshold energy densityfor an antimicrobial and/or antifungal phenomenon based on the uniquewavelengths of energy delivery being simultaneous (λ=870 nm and 930 nmtogether), or using a 930 nm wavelength alone.

Therefore, it is desirable to NIMELS antimicrobial therapy that thismethod of (Energy Density) quantification is conserved and the novelvalue of the NIMELS Factor (Tn) is used to calculate the necessary,parabolic reciprocal correlations for safe and effective dosimetryvalues.

This NIMEL method of temporal and reciprocal dosimetry should also holdtrue for differences in laser output power (between 1 W -5 W) as long asany quantifiable thermal increase, thermal increase time durations, andphotobiological events in the tissues are kept below any irreversibledamage threshold values.

EXAMPLE XII NIMELS Therapy used with Exemplary Medical Devices

The following examples, described with particular reference to FIGS.16-22, are provided to illustrate the application of the NIMELtechnology in the medical devices area as discussed above. Accordingly,the embodiments described are provided as representative examples. Oneof skill will appreciate that multiple permutations and variationsexploiting the underlying NIMEL methodology may be devised.

FIG. 16 shows an embodiment of a NIMELS Optical Catheter Controllerincluding delivery assembly configured as multiple optical fibersembedded into a catheter controller around a catheter entry port placedon a patient. FIG. 17 shows a physical model constructed to simulate theembodiment of FIG. 16.

FIGS. 16 and 17 show a connectable adapter whereby a spray of opticalfibers are imbedded within a disposable percutaneous device controller,and distally connected to a NIMEL laser system. In many differentvariations depending on the size of the percutaneous device, a pluralityof optical fiber sprays are imbedded in a circular (or other)overlapping pattern, to enable irradiation on the percutaneous wound forthe percutaneous device. According to this embodiment of the invention,the fibers are bundled together at one end, where they can be connectedto NIMELS laser system, and at the other end unrestrained to flareoutwardly forming a spray, to embed in a necessary pattern in thepercutaneous device controller bandage.

FIG. 18 depicts the underside of a NIMELS Optical Catheter Controllersimilar to FIG. 16. FIG. 19 shows a physical model according to FIG. 18,with the optical fibers removed.

FIGS. 18 and 19 illustrate the illumination of optical fiber arrays forthe irradiation of percutaneous wounds with percutaneous devicecontrollers. The adapter carries the bundled end of the optical fibersat one end, and is formed to engage an adapter connected to a NIMELSlaser system. The fiber optic can transmit NIMELS energy to a varietyand plurality of different locations throughout the percutaneous devicecontroller, and the percutaneous device itself. The fiber optic cablecan include a plurality of optical fibers, each of which individuallyterminates at one of a plurality of sites in and around the percutaneousdevice controller and the percutaneous device itself. This can include astepped or Bragg graded fiber for the internal lumen irradiation of apercutaneous device. Alternatively, the optical fibers can individuallyterminate at desired, e.g., evenly spaced, locations throughout thedevice to illuminate a region of the percutaneous device controlleruniformly.

FIG. 20 is prototype enabled side view of a NIMELS Optical MicrobialCatheter Controller according to the present disclosure. FIG. 21 is anadditional view of the prototype of FIG. 20.

FIGS. 20 and 21 illustrate a Radiation Dispersion Bandage or OpticalPercutaneous Device Controller for use as an adjunctive treatment for aninfected percutaneous device or to prevent infection and colonization ofa percutaneous device. The device can alternatively include flexibleilluminators for the external and internal phototherapy of apercutaneous device controller and/or percutaneous device itself. Theilluminators may be formed so as to be imbedded or wrapped in or arounda percutaneous device controller and or percutaneous device itself. Inanother configuration, the illuminators may be actively or passivelycooled so the percutaneous wound, skin, and or device itself remainsbelow a desired temperature.

Also, a flexible band or belt may be provided with the percutaneousdevice controller to permit the device to be held or contoured to adesired body surface for the adequate positioning and illumination ofthe percutaneous device. The Optical Percutaneous Device Controller canbe designed (e.g., configured and arranged) to swathe tightly aroundvascular and non-vascular percutaneous devices, providing extendedantimicrobial environments (with NIMELS energy) for extended periods oftime.

FIG. 22 is a further view of a NIMELS Optical Microbial CatheterController according to the present disclosure.

As noted previously, a delivery assembly used according to the presentdisclosure may take forms other than optical fibers. For example, hollowwaveguides may be used for the delivery assembly in certain embodiments.Other size and shapes for the deliver assembly, e.g., assembly 14 inFIG. 2, may also be employed based on the requirements of theapplication site. In exemplary embodiments, the delivery assembly 14 canbe configured for free space or free beam application of the opticalradiation, e.g., making use of available transmission through tissue atNIMELS wavelengths described herein. For example, at 930 nm (and to asimilar degree, 870 nm), the applied optical radiation can penetratepatient tissue by up to 1 cm or more. Such embodiments may beparticularly well suited for use with in vivo medical devices asdescribed below. Suitable collimating and/or aperture stop opticalelements may be used.

Accordingly, applications of NIMELS techniques of the present disclosurecan be used with medical devices including, the but not limited to, IVCatheters, such as PICC sites, Central Venous (CV) Lines, ArterialCatheter, Peripheral Catheters, Dialysis Catheters, External fixatorpins, Peritoneal dialysis catheters, Epidural catheters, Chest tubes,Gastronomy feeding tubes as illustrated in FIG. 13.

EXAMPLE XIII In Vitro Safety Testing—Mammalian Cells

Conventional mouse 3T3 fibroblasts were used to determine whethermammalian cells were injured by the NIMELS laser treatment. Treatmentplates containing a standardized quantity of fibroblasts were exposed tothe NIMELS laser; control plates containing the same quantity offibroblasts were held at room temperature for the duration of lasertreatment.

After treatment, the cells were allowed to attach to their plates forthree hours in a 37° C. incubator. The cells were then extracted fromthe plates and examined for morphology and viability. While there weremorphological changes observed in the treated fibroblasts, the viabilityof the treated and control plates showed no significant difference.These results indicate that any cell damage (as demonstrated bymorphological changes) did not affect cellular viability.

An additional in vitro study was undertaken to test for thermal andoptical safety to mouse 3T3 fibroblast tissue when exposed to NIMELSlaser dosimetry shown to be lethal to bacteria in vitro. Fibroblastcells were inoculated with 500,000 CFU of E. coli K-12. This “infected”sample was treated with the microbial-lethal laser dosimetry ascertainedfrom prior studies (see, supra). Three hours after treatment, thesefibroblast cells appeared viable in shape and morphology. In culturesperformed at 16 hours post-treatment, there was no bacterial growth instandard agar and mammalian growth serum medium.

EXAMPLE XIV In Vivo Safety Testing—Mammalian Cells

Based on the in vitro results, a study was performed in mice todetermine the safety of the NIMELS laser at these wavelengths in ananimal model.

A pilot dosimetry study was conducted using a NIMELS laser on the dorsalskin of the FVB (Friend leukemia virus B strain) mouse strain. Sixgroups of four mice each were used. This included the testing of laserintensity, energy level, power density (PD), exposure time and spotsize. Observations were made on the mice on the day of study (day 0),with follow up observations conducted on day 1 and 2. The mice weresacrificed on day 2 and sections from the laser-exposed region wereprepared for histological examination by paraffin embedding followed byHematoxylin and Eosin (H&E) staining.

All animal deaths, serious morbidity and visible scarring of the skinoccurred in animals where energy (range 888-3034 J), power density(range 2.04-3.82) levels or exposure time were used that far exceededthose contemplated above for use with mammals according to theinvention.

Thirty-four (34) specimens were studied microscopically. All came fromanimals that survived the initial treatment and lived through theobservation period. The histology revealed that 28 of the 34 specimensshowed no histological abnormality, of which 6 were controls, and 22 hadbeen exposed to laser energies ranging from 360 J to 1776 J and PDranging from 1.02 to 2.72.

There were six specimens that demonstrated positive histology, three ofwhich had been exposed to laser energies far greater than 750 J (rangingfrom 1332 J to 1998 J). Of the remainder, one had been exposed toextremely high power density (PD 3.82, 444 J) and one was subjected tosignificantly prolonged exposure time (4 minute exposure with 930 nm,750 J). In the remaining specimen, the exposure factors were within therange anticipated for human use.

The intensity of histological changes was carefully noted. It isnoteworthy that in none of the specimens in which there werehistological changes noted, even those subjected to extreme exposures,none of the findings extended subcutaneously to the underlying muscularlayer or beyond. The changes were extremely superficial, penetrating toa depth of less than 90 microns. The surface ulcerations noted wererecognizable only after careful microscopic search and were less than 60microns in diameter and 40 microns in depth. Therefore, all changes wereconsidered minor and of little clinical consequence.

A strong correlation with adverse events was noted with energy levelsused (Joules), where all events increase in number and intensity as theJ level increased. With one exception, no major adverse outcomes werenoted when laser energy of 750 J or less was used. In only one animalwas there some evidence of skin scarring noted at 750 J and thatoccurred when the duration of the laser exposure was extended to twicethe usual time anticipated for use in mammals (e.g., humans).

The study revealed that there was no apparent damage to the skin orunderlying tissue in the majority of the animals. Severe morbidity wasconfined to the animals in which very high energy levels were employed.The number and intensity of all serious adverse outcomes related to theintensity of the exposure, all occurring when physical parameters wereused that far exceeded anticipated human use.

Positive histological findings were encountered across a broad range ofparameters, and were clearly more prominent when higher energy levels orhigher power density was employed. Most importantly, where levels wereemployed that were within the range of anticipated use in mammals, thefindings were either normal or, if abnormal, were of an extremely minornature, recognizable only after extensive microscopic search.

The study underscores the necessity of not only monitoring the energylevels (J) employed, but the necessity of maintaining proper control ofexposure time and laser beam spot size, each of which will significantlyaffect power density (PD).

EXAMPLE XV In Vivo Safety Testing—Human Patient

Following the in vitro fibroblast studies, the inventor performed adosimetry titration on himself to ascertain the safe, maximum level ofenergy and time of exposure that could be delivered to human dermaltissue without burning or otherwise damaging the irradiated tissues.

The methodology he used was to irradiate his great toe for varyinglengths of time and power settings with the NIMELS laser. The results ofthis self-exposure experiment are described below.

TABLE XXII Combined Wavelength Dosimetries OUTPUT BEAM TOTAL ENERGYPOWER POWER SPOT AREA OF TIME ENERGY DENSITY DENSITY PARAMETERS (W) (CM)SPOT (CM²) (SEC) JOULES (J/CM²) (W/CM²) 870 nm 1.5 1.5 1.77 250 375 2120.85 930 nm 1.5 1.5 1.77 250 375 212 0.85 Combined 3.0 1.5 1.77 250 750424 1.70

TABLE XXIII Dosimetry at λ = 930 nm OUTPUT BEAM TOTAL ENERGY POWER POWERSPOT AREA OF TIME ENERGY DENSITY DENSITY PARAMETERS (W) (CM) SPOT (CM2)(SEC) JOULES (J/CM²) (W/CM²) 930 nm 3.0 1.5 1.77 120 360 204 1.70

Time/Temperature assessments were charted to ensure the thermal safetyof these laser energies on human dermal tissues (data not shown). In onelaser procedure, he exposed his great toe to both 870 nm and 930 nm forup to 233 seconds, while measuring toenail surface temperature with alaser infrared thermometer. He found that using the above dosimetries,at a surface temperature of 37.5° C., with 870 nm and 930 nm togetherwith a combined Power Density of 1.70 W/cm², pain resulted and the laserwas turned off.

In a second laser procedure, he exposed his great toe to 930 nm for upto 142 seconds, while again measuring toenail surface temperature with alaser infrared thermometer. He found that, at a surface temperature of36° C., with 930 nm alone at a Power Density of 1.70 W/cm², painresulted and the laser was turned off.

EXAMPLE XVI In Vivo Safety Testing—Limited Clinical Pilot Study

Following the experiment above, additional patients with onychomycosisof the feet were treated. These patients were all unpaid volunteers, whoprovided signed informed consent. The principle goal of this limitedpilot study was to achieve the same level of fungal decontamination invivo, as was obtained in vitro with the NIMELS laser device. We alsodecided to apply the maximum time exposure and temperature limit thatwas tolerated by the inventor during his self-exposure experiment.

In a highly controlled and monitored environment, three to five laserexposure procedures were performed on each subject. Four subjects wererecruited and underwent the treatment. Subjects provided signed informedconsent, were not compensated, and were informed they could withdraw atany time, even during a procedure.

The dosimetry that was used for the treatment of the first subject wasthe same as that used during the inventor's self-exposure (shown above).The temperature parameters on the surface of the nail also wereequivalent to the temperatures found by the inventor on self-exposure.

The treated toes showed significantly reduced Tinea pedis and scalingsurrounding the nail beds, which indicated a decontamination of the nailplate that was acting as a reservoir for the fungus. The control nailswere scraped with a cross-cut tissue bur, and the shavings were saved tobe plated on mycological media. The treated nails were scraped andplated in the exact same manner.

For culturing the nail scrapings, Sabouraud dextrose agar (2% dextrose)medium was prepared with the following additions: chloramphenicol (0.04mg/ml), for general fungal testing; chloramphenicol (0.04 mg/ml) andcycloheximide (0.4g/ml), which is selective for dermatophytes;chloramphenicol (0.04 mg/ml) and griseofulvin (20 μg/ml), which servedas a negative control for fungal growth.

Nine-day mycological results for Treatment #1 and Treatment #2(performed three days after Treatment #1) were the same, with adermatophyte growing on the control toenail plates, and no growth on thetreated toenail plates. Treated plates did not show any growth whereasuntreated control culture plates showed significant growth.

The first subject was followed for 120 days, and received fourtreatments under the same protocol. FIG. 15 shows a comparison of thepretreatment, 60 days post-treatment and 80 days post-treatment, and 120days post-treatment toenails. Notably, healthy and non-infected nailplate was covering 50% of the nail area and growing from healthy cuticleafter 120 days.

While certain embodiments have been described herein, it will beunderstood by one skilled in the art that the methods, systems, andapparatus of the present disclosure may be embodied in other specificforms without departing from the spirit thereof. The present embodimentsare therefore to be considered in all respects as illustrative and notrestrictive of the present disclosure.

1. A method of reducing the level of a biological contaminant in atarget site without an intolerable adverse effect on a biologicalmoiety, comprising the step of irradiating the target site with anoptical radiation having a first wavelength from about 865 nm to about875 nm, or a second wavelength from about 925 nm to about 935 nm, orboth wavelengths, and at a NIMELS dosimetry.
 2. The method of claim 1,wherein the biological contaminant comprises bacteria, fungi, or molds.3. The method of claim 2, wherein the biological contaminant is aselected from the group consisting of Trichophyton, Microsporum,Epidermophyton, Candida, Scopulariopsis brevicaulis, Fusarium spp.,Aspergillus spp., Alternaria, Acremonium, Scytalidinum dimidiatum, andScytalidinium hyalinum.
 4. The method of claims 1, wherein thebiological contaminant is E. coli.
 5. The method of claim 1, wherein thebiological contaminant is Staphylococcus or MRSA.
 6. The method of claim1, wherein the optical radiation is provided for a time (Tn) of fromabout 50 to about 450 seconds.
 7. The method of claim 1, wherein theNIMELS dosimetry provides an energy density from about 200 J/cm² toabout 700 J/cm².
 8. The method of claim 1, wherein the NIMELS dosimetryprovides an energy density from about 275 J/cm² to about 500 J/cm².
 9. Atherapeutic system comprising: an optical radiation generation deviceconfigured and arranged to generate optical radiation substantially in afirst wavelength range from about 865 nm to about 875 nm or a secondoptical radiation range having a wavelength from about 925 nm to about935nm, or both wavelength ranges; a delivery assembly for causing theoptical radiation to be transmitted through an application region; and acontroller operatively connected to the optical radiation generationdevice for controlling dosage of the radiation transmitted through theapplication region at a NIMELS dosimetry, whereby the time integral ofthe power density of the transmitted radiation per unit area is below apredetermined threshold of a NIMELS dosimetry.
 10. A therapeutic systemaccording to claim 9, wherein the optical radiation source includes adiode laser configured and arranged to produce an output in the nearinfrared region to be delivered to one or more anatomical regionssimultaneously.
 11. A therapeutic system according to claim 9, whereinthe controller is configured and arranged to control the radiation to bean output of a succession of radiation pulses or a continuous wave. 12.A therapeutic system according to claim 9, wherein the controllercomprises a dosimetry calculator which is preprogrammed to calculatedosage needed for treatment of the treatment site.
 13. A therapeuticsystem according to claim 12, wherein the dosimetry calculator furthercomprises an imaging system for imaging the treatment site andgenerating the anatomic data of the treatment site.
 14. A therapeuticsystem according to claim 9, wherein the delivery assembly comprises oneor more optical fibers configured and arranged to receive radiation fromthe optical radiation generation device and deliver the radiation to amedical device disposed in tissue of a patient (in vivo).
 15. Atherapeutic system according to claim 14, wherein the medical device isa stent.
 16. A therapeutic system according to claim 14, wherein themedical device is an artificial joint.
 17. A therapeutic systemaccording to claim 14, wherein the medical device is a catheter.
 18. Atherapeutic system according to claim 14, wherein the medical device isselected from the group consisting of an IV catheter, a central venousline, an arterial catheter, a peripheral catheter, a dialysis catheter,an external fixator pin, peritoneal dialysis catheter, an epiduralcatheter, a chest tube, and a gastronomy feeding tube.
 19. Thetherapeutic system of claim 9, wherein the optical radiation is providedfor a time (Tn) of from about 50 to about 450 seconds.
 20. Thetherapeutic system of claim 9, wherein the NIMELS dosimetry provides anenergy density from about 200 J/cm² to about 700 J/cm².